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The University of East Anglia at the Ramada Hotel, Norwich

19th – 23rd April 2009

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Contents

Conference Programme 2

General Information for Delegates 6

Sponsors 9

Committee of the ESR Group of the RSC 10

The Henry Wellcome Unit at UEA 11

JEOL student Prize Lectures and Reception 12

Bruker Lecture and Reception 13

Next Meeting – Cardiff 2010 14

Announcements 15

Abstracts for Talks 17

Abstracts for Posters 62

Title Index 96

Author Index 101

List of Participants 103

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Conference Programme All Conference Lectures will be held in the City Suite at the Ramada Hotel. The poster session will take place at the City Suite Reception Area/Lounge. SUNDAY 19th APRIL

09:00 – 17:00 COST P15 Training School

Club Room, Ramada Hotel

SUNDAY 19th APRIL

- - - - - - - - - - - - Arrival - - - - - - - - - - - - 14:00 Check-in opens Foyer, Ramada Hotel

16:30 - 18:00 Registration opens 2nd Avenue, Ramada Hotel

18:00 Bar opens Presidential Suite Bar

18:30 Dinner Presidential Suite

20:00 RSC Wine Reception

City Suite Reception Area

MONDAY 20th APRIL

07:30 – 08:50 Breakfast Arts Restaurant

08:00 – 09:00 Registration open

Session Chair: Professor David Collison

08:55 – 09:00 City Suite Conference opening and welcome note

09:00 – 09:40 Wayne L. Hubbell Keynote Lecture: Functional protein dynamics from site-directed spin labelling

09:45 – 10:00 Sergei Dzuba PELDOR studies of conformations of single- and double-stranded DNA with non-nucleotide inserts

10:05 – 10:20 Jeffrey Harmer Orientation selective DEER measurements in systems containing paramagnetic iron-sulfur clusters

10:30 – 11:00 Tea & Coffee City Suite Reception Area

Session Chair: Dr Fraser MacMillan

11:00 – 11:25 R. David Britt Invited Lecture: Multifrequency Pulsed EPR targeting metal cluster - ligand interactions

11:30 – 11:45 Bruno Guigliarelli Engineering dioxygen sensitivity of hydrogenases: Mechanistic studies combining EPR, photochemistry and electrochemistry

11:50 – 12:05 Alistair Fielding Multifrequency Electron Paramagnetic Study of the Radical Intermediates Generated by PpoA, a Novel CYP450 Fusion Protein from Aspergillus nidulans

12:10 – 12:25 Stefan Stoll Substrate radical intermediates in cyanobacterial bilin reductases

12:30 – 13:45 Lunch Arts Restaurant

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Session Chair: Professor Andrew J. Thomson, FRS, OBE

14:00 – 14:15 Angelika Boeer JEOL Student Prize Talk: Anisotropy in Molecular Magnetism; Magnetic Exchange Coupling of Octahedral Cobalt(II) Ions

14:20 – 14:35 Susanna Pudollek JEOL Student Prize Talk: EPR and 55Mn-ENDOR Spectroscopy of the S2-state Multiline Signal of Photosystem II

14:40 – 14:55 Bela Bode JEOL Student Prize Talk: Exotica in pulsed dipolar spectroscopy; cobalt(II)-nitroxide distances and spatial distribution of nitroxides in membranes and detergents

15:00 – 16:20 Tea & Coffee / "odd" Posters

City Suite Reception Area

Session Chair: Professor Martin Kaupp

16:20 – 16:45 David Keeble Invited Lecture: Doping of Perovskite Oxide Functional Materials: Local structure determination using Electron Magnetic Resonance

16:50 – 17:05 Sepideh Zamani EPR analysis of Vanadium-doped mesoporous silica and titania catalysts

17:10 – 17:25 Stephen Sproules An "EQR" spectrum: Electronic structure of rhenium tris(dithiolene) electron transfer series

17:30 – 17:45 Chandrima Pal CW and Pulse ESR study on partially zinc (II) exchanged and partially dehydrated zeolite

18:00 Bar opens Presidential Suite Bar

18:30 Dinner Presidential Suite

20:00 JEOL Reception / Poster Viewing

City Suite Reception Area

TUESDAY 21st APRIL

07:30 – 08:50 Breakfast Arts Restaurant

Session Chair: Professor R. David Britt

09:00 – 09:40 Fraser MacMillan Keynote Lecture: Correlating the structure & function of proteins using EPR

09:45 – 10:00 Janet Lovett DEER; Probing nanosized self-assembled structures

10:05 – 10:20 Enrica Bordignon Reciprocal transmembrane signaling in an ABC transporter

10:30 – 11:00 Tea & Coffee City Suite Reception Area

Session Chair: Professor Richard Cammack

11:00 – 11:25 Inés García Rubio Invited Lecture: Tuning redox activity by replacing the proximal heme ligand in cytochrome P450cam with selenocysteine

11:30 – 11:45 Christian Teutloff The electronic structure and protein interactions of YD in PS-II investigated by high-field ENDOR-spectroscopy

11:50 – 12:05 Nikolay Isaev Fast Stochastic Librations and Slow Rotations of Spin Labeled Stearic Acids in a Model Phospholipid Bilayer at Cryogenic Temperatures

12:10 – 12:25 David Norman Using PELDOR to define and refine the domain orientation of modular proteins

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12:30 – 13:45 Lunch Arts Restaurant

13:45 – 17.45

Free time

Bus transfer into Norwich & Tour of Norwich Cathedral

18:00 Bar opens Presidential Suite Bar

18:30 Dinner Presidential Suite

Session Chair: Professor David Collison

20:00 Gunnar Jeschke Bruker Lecture: Measuring the Nanoworld

21:00 Bruker Reception / Poster Viewing

City Suite Reception Area

WEDNESDAY 22nd APRIL

07:30 – 08:50 Breakfast Arts Restaurant

Session Chair: Dr. Christopher Kay

09:00 – 09:40 Martin Kaupp Keynote Lecture: Quantum Chemical Calculations of EPR Parameters as a Tool to Study Metalloenzyme Sites Sponsored by the EPSRC National Service for EPR Spectroscopy

09:45 – 10:00 Ilya Kuprov State space reduction and symmetry factorization in EPR and Spin Chemistry simulations

10:05 – 10:20 Dimitri Svistunenko Revisiting the McConnell relationship: why C1?

10:30 – 11:00 Tea & Coffee City Suite Reception Area

Session Chair: Professor Gunnar Jeschke

11:00 – 11:25 Marina Bennati Invited Lecture: 1H and 13C Dynamic nuclear polarisation with a two-field (0.35/14T)shuttle DNP spectrometer

11:30 – 11:45 Graham Smith Very high bandwidth, orientation selective, DEER spectroscopy at 94GHz

11:50 – 12:05 Henk Vrielinck ENDOR in field-frequency space: orientation, species and quantum state selection

12:10 – 12:25 Hideo Utsumi Development of high field OMRI scanner for imaging in vivo redox status in mouse

12:30 – 13:45 Lunch Arts Restaurant

Session Chair: Dr. Victor Chechik

14:00 – 14:15 Philipp Neumann Single spins in diamond: quantum computing and magnetometry

14:20 – 14:35 Mark Newton Quantum tunnelling of hydrogen in the nitrogen; vacancy; hydrogen complex in diamond

14:40 – 14:55 Mohamed Morsy An EPR Structural and Conformational Evidences of Liquid Crystalline and Carbon Nanotube Systems using Different Radical Probes

15:00 – 15:15 Christopher Smith ESR study on Hydrogen Absorption property of Ball-milled Graphite

15:20 – 16:20 Tea & Coffee / "even" Posters

City Suite Reception Area

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Session Chair: Dr. Mark Newton

16:20 – 16:35 Christopher Wedge Time-Resolved Low-Field EPR of a Carotenoid-Porphyrin-Fullerene Triad

16:40 – 16:55 Iain McKenzie Probing Soft Matter with Avoided Level Crossing Muon Spin Resonance: A Complimentary Technique to EPR

17:00 – 17:15 Katharina Pirker Reactions of Cu(II) with teas and polyphenols

17:20 – 17:35 Rachel Haywood Ascorbate (vitamin C) accelerates hydrogen-peroxide-induced melanoma cell death via modulation of oxidative stress: vitamin C a DNA damage switch?

17:45 RSC ESR Group AGM of the RSC ESR Group (all welcome to attend)

17:45 – 19:00 Poster Viewing City Suite Reception Area

19:30 Pre-dinner drinks City Suite Reception Area

20:00 Banquet City Suite THURSDAY 23rd APRIL

07:30 – 08:50 Breakfast Arts Restaurant

Check-out of Hotel rooms by 13.00 (luggage storage will be available)

Session Chair: Dr. Rachel Haywood

09:00 – 09:40 Richard Cammack Keynote Lecture: Biochemical targets for advanced EPR weaponry

09:45 – 10:00 Georg Gescheidt Investigations on Radical Polymerisation by CIDNP; New Insights

10:05 – 10:20 Kiminori Maeda Spin dynamics of the model system of a chemical compass using pulsed EPR and transient absorption spectroscopies

10:30 – 11:00 Tea & Coffee City Suite Reception Area

Session Chair: Dr. Damien Murphy

11:00 – 11:15 Sergey Semenov Host-guest complexes of pH-sensitive nitroxides for EPR spectroscopy and imaging

11:20 – 11:35 Johann Klare Structure and function of the tRNA modifying MnmE/GidA complex studied with DEER

11:40 – 11:55 Olav Schiemann PELDOR on the 320kD Capsular Export Channel Wza

12:00 – 12:15 Victor Chechik Au nanoparticle catalysed oxidation reactions: a spin trapping study

12:30 – 13:45 Lunch Arts Restaurant

- - - - - - - - - - - - Departure - - - - - - - - - - - - Additional Meetings Sunday 20.45 – 21.30 Committee Meeting of the ESR Group (Statesman Room) Tuesday 14.00 – 16.00 Oxford SAC Meeting (Statesman Room) Tuesday 16.00 – 18.00 Manchester MAP Meeting (Statesman Room) Wednesday 17.45 – 18.30 Annual General Meeting of the ESR Group (City Suite)

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General Information All activities will take place at the Ramada Hotel, Norwich (see Transport section).

Registration The hotel check-in time is from 2pm onwards (Hotel reception is open 24 hours). Registration will take place on the Sunday evening from 16:30 till 18:00 and on Monday morning from 08:00 till 09:00 at the Registration Desk which can be found on 2nd Avenue just across from the main Hotel reception (see signposts).

Lectures All lectures will be held in the City Suite.

Poster Sessions Posters will be located in the City Suite Reception Area (at the rear of the City Suite) and should be displayed for the duration of the conference. There will be two poster sessions held on the Monday ("ODD") and Wednesday ("EVEN") afternoons. Please hang your posters on Sunday evening and remove them on Thursday morning.

Tea and Coffee Breaks Tea and coffee breaks will be served in the City Suite Reception Area. Iced water and tea/coffee will be available at all times during the sessions.

Lunches and Dinners Lunches will be at 12:30 until 13:45 in the Arts restaurant and evening meals (apart from the banquet) will be at 18:30 each day in the Presidential Suite (basement). The Presidential Suite bar will open at 18:00 each day. The banquet will be held in the City Suite at 20:00 on the Wednesday.

Accompanying persons We do not offer a specific programme for accompanying persons, but there are many sites of interest to visit. For information, please ask any of the local organisers or see information displayed at the Registration desk.

Information for speakers Please allow 5 minutes for discussion at the end of all lectures. Excluding discussion time, keynote lectures will be 40 minutes, invited lectures 25 minutes, JEOL Student Prize talks and contributed lectures 15 minutes. Speakers are requested to upload their presentation a day prior to their talk at the latest. Please hand in your file (pdf or ppt) to the Registration desk or to a local organiser (red button on badge). Make sure you enable the

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“Embed all fonts” option when saving your presentation to avoid font and figure corruption. Please would all speakers ensure they keep strictly to the time schedule for their talks. Chairs are also advised to adhere to their session time schedule.

Suggestions for Tuesday Afternoon Tour of Norwich Norman Cathedral The magnificent Cathedral dominates the city skyline. Situated in the heart of the city, Norwich Cathedral has attracted many pilgrims and visitors for over 900 years. Separated from the busy streets by flint walls and entrance gates, it is a place of great splendour and tranquillity and has at least three services daily. The Cathedral was the vision of Herbert de Losinga, first Bishop of Norwich and construction commenced in 1096. However, the Cathedral was not finally consecrated until 1278, built mainly of Caen stone, a pale honey-coloured limestone brought over from Normandy, with Norfolk flints and stone from Northamptonshire. The Cathedral spire is 315 feet (96 m) high, the second highest in England and with the largest monastic cloister. It is also one of the finest complete Romanesque buildings in Europe. On Tuesday afternoon we have arranged for a guided tour of the cathedral including coach transport to and from the Hotel. Numbers are limited and we ask that you sign up (on a first-come, first-served basis) for this event during registration or by Monday lunchtime at the latest.

Shopping in Norwich Norwich is a unique shopping destination with its large pedestrian areas, quirky, independent shops, 6 day open-air market and spacious modern shopping malls. Range, quality and that element of surprise are all part of the Norwich shopping experience. The city’s “High Street” includes St Stephen’s and pedestrianised thoroughfares like Gentleman’s Walk and London Street. The Mall Norwich and Chapelfield shopping centre have over 150 stores between them and, together with the city centre, Norwich has all the UK’s leading department stores, including John Lewis, Marks & Spencer, House of Fraser, Bhs and Debenhams. Dozens of independent shops bring an extra dimension to any trip. Jarrold’s has been named UK Independent Department Store of the Year twice and the Norwich Lanes and Timberhill offer individual fashion and lifestyle shopping, against a background of narrow alleys and beautiful historic buildings. Elm Hill, Norwich’s most famous medieval cobbled street, combines beautifully preserved timber framed houses with shops selling everything from antiques to teddy bears!

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Transport to and around Norwich Getting to Norwich By Road: Major trunk roads to the Norwich area are the M11, A11, A12, A140 (from London, the south, the ferry ports and the Channel Tunnel) and the A14, A47, A1/M1 (from the north and west). By Train: Norwich Railway Station is located in central Norwich, a 5 minute walk from the city centre. Train operator National Express East Anglia runs trains to Norwich from London's Liverpool Street Station, every 30 minutes during the day. The journey time is approximately 1 hour 50 minutes. A direct service also links Cambridge to Norwich, with an approximate journey time of 1 hour 10 minutes. Central Trains offer connecting services from the Midlands, north of England and Scotland via Peterborough. Details of local rail travel and on-line train timetables are available (www.nationalrail.co.uk) or if you prefer to phone contact National Rail Enquiries on 08457 48 49 50. Please Note! There are special arrangements for travel on Sunday April 19th due to engineering works between London and Norwich and trains will be replaced by buses for part of the journey extending the journey time considerably. An alternative option is to travel via Cambridge and Ely departing either from London Liverpool Street or London Kings Cross. Participants should look at www.nationalrail.co.uk/index.html to plan their route. By Coach: National Express coaches travel to Norwich from London and other major cities several times daily. The London to Norwich journey time is around 3 hours. National Express Airport links the main London airports to Norwich with a regular direct service - up to 10 times a day from Stansted, Heathrow and Gatwick airports. For fares and times: 08705 80 80 80 or www.nationalexpress.com By Air: Norwich is home to a major airport, Norwich International, which is just 4 miles from the city centre, serving many UK and international destinations (www.norwichairport.co.uk). The 42nd Annual International Meeting of the ESR Group of the RSC will be held at the Ramada Hotel in Norwich, UK. The site is located on the Norwich outer ring road, and is only about 1 mile from Norwich's International airport. For further details, visit www.ramadajarvis.co.uk/hotels/norwich/travel.aspx.

Transport around Norwich The hotel has a mini-bus service which can be used. On the Tuesday afternoon there will be two buses running into the city and back for those interested. Taxi numbers, if required, are e.g. (01603) 666333 / (01603) 455555; others are available at the hotel reception desk.

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Conference Sponsors: We are grateful to our sponsors: The University of East Anglia (Faculty of Science), Bruker Biospin, JEOL UK, The Royal Society of Chemistry, COST P15, Ramada Hotels, œrlikon, Photonic Solutions, Goss Scientific, barthel HF-Technik, Mathworks, SpringerWienNewYork and Taylor & Francis. We encourage participants to view the sponsor displays during the meeting.

(www.photonicsolutions.co.uk)

(www.oerlikon.com)

(www.barthel-hf.com)

(www.rsc.org)

(www.ueac.ac.uk/sci)

(www.jeoluk.com)

(www.bruker.com/uk.html)

(www.springer.com/springerwiennewyork)

(www.taylorandfrancis.com)

(www.mathworks.co.uk)

(www.ramadajarvis.co.uk/hotels/norwich)

(www.st-andrews.ac.uk/~costp15) (www.gossinst.com)

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Committee of the ESR Spectroscopy Group of the Royal Society of Chemistry Prof David Collison (Chairman) University of Manchester

Dr Chris Kay (Secretary) University College London

Dr Victor Chechik (Treasurer) University of York

Dr Fraser MacMillan (Local Organiser) University of East Anglia

Dr. Rachel Haywood RAFT Institute

Dr Helen Williams AstraZeneca

Dr. Damien Murphy University of Cardiff

Dr Mark Newton University of Warwick

Dr Sean McWhinnie Royal Society of Chemistry

Dr Louise Ottignon (Companies Representative) Bruker Biospin

Local Organising Committee (all University of East Anglia)

Dr Fraser MacMillan (Local Chairman)

Matthew Bawn

Dr. Justin Bradley

Dr. Myles Cheesman

Professor Andrew J. Thomson

Dr. Jessica van Wonderen

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Local Organisation at UEA The School of Chemical Sciences and Pharmacy (CAP)

Chemistry research within the School of Chemical Sciences & Pharmacy of which biophysical chemistry is a major theme, is led by 28 faculty members. There are currently about 500 undergraduate students, about 100 graduate students studying for MSc and PhD degrees and about 40 post-doctoral workers. The School's current research grant holding is in excess of £11M. Over the past 3 years more than £9M has been awarded to the School to provide for laboratory refurbishment and the acquisition of major new instrumentation, including a JIF grant of £3.6M for the further development of biophysical chemistry research, over £5.5M from the Wolfson Foundation, HEFCE, the Weston Foundation and SRIF for laboratory refurbishment and the development of new research programmes. Institutes affiliated to the University include the internationally renowned John Innes Centre, Sainsbury Laboratory, and Institute of Food Research. More information about can be found at http://www.uea.ac.uk/cap

Centre for Molecular and Structural Biochemistry The UEA Centre for Metalloprotein Spectroscopy and Biology (CMSB) was originally formed from faculty members in biological and chemical sciences at UEA with the broad aim of understanding: (i) the activation and redox cycling of inorganic substrates involved in the bacterial nitrogen, oxygen and sulphur cycles and (ii) metal ion metabolism and metal-microbe interactions. Since its inception 15 years ago the CMSB has developed an international reputation for excellence in its field based on publication of over 180 papers in peer reviewed journals. Moreover the centre is highly regarded as an exemplar of how to successfully organise science at the life science: chemistry interface. Over the years, members of the CMSB have found themselves no longer working exclusively on redox-active metalloproteins and to reflect this they have recently rebranded CMSB as the Centre for Molecular and Structural Biochemistry under the directorship of Prof. David Richardson. The rebranded CMSB was launched in May 2008. The expertise and techniques within the CMSB comprise microbial energetics, molecular biology, fermentation and large scale protein preparation, flow-flash and stopped flow kinetics, paramagnetic spectroscopies including magnetic circular dichroism and electron paramagnetic resonance spectroscopies, high field nuclear magnetic resonance spectroscopy, thin film protein electrochemistry and X-ray crystallography.

The Henry Wellcome Unit for Biological EPR at UEA The Henry Wellcome Unit for Biological EPR, established at UEA in 2002 by the Wellcome Trust/HEFCE Joint Infrastructure Fund (JIF), houses a range of state-of-the art EPR equipment including the UK's first Bruker ELEXSYS E680 FT Hybrid pulsed EPR instrument operating at two frequencies, 9 GHz (X-band) and 95 GHz (W-band), in both continuous wave (CW) and pulsed modes. It is fitted with both PELDOR and ENDOR accessories. In addition, there is a CW X-band ELEXSYS instrument with low loss cavity for measurement of aqueous samples at room temperature as well as a further multi-frequency instrument (at S-, X- and Q-band). Thus UEA has one of the best equipped EPR laboratories in the UK and is experienced in undertaking advanced EPR studies of membrane proteins.

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JEOL Student Prize Lectures and Reception The JEOL student lecture competition started at the Lancaster University meeting and is now in its 14th year. The competition is open to postgraduates in their 2nd or 3rd year and postdoctoral fellows in their 1st year. The 15 minute lectures are judged by the ESR Group Committee on the basis of their scientific content and delivery. An engraved medal and monetary prize are kindly provided by JEOL for the winner of the competition, to be presented at the conference banquet. This year the competition will take place in the City Suite during the Monday afternoon session. The 2009 lectures, selected on the basis of the abstracts submitted, will be:

Anisotropy in Molecular Magnetism; Magnetic Exchange Coupling of Octahedral Cobalt(II) Ions

Angelika Boeer, University of Manchester, UK

EPR and 55Mn-ENDOR Spectroscopy of the S2-state Multi-line Signal of Photosystem II

Susanne Pudollek, Freie Universität Berlin, Germany

Exotica in pulsed dipolar spectroscopy: cobalt(II)-nitroxide distances and spatial distribution of nitroxides

in membranes and detergents

Dr. Bela Bode, J. W. Goethe Universität Frankfurt, Germany The wine at dinner on Monday evening and refreshments at the subsequent wine reception during the evening Poster session are sponsored by JEOL.

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Bruker Lecture and Reception Since 1986 Bruker BioSpin has generously sponsored an annual lectureship and prize, given to a scientist who has made major contributions to the application of ESR spectroscopy in chemical or biological systems. The Bruker Lectureship for 2009 has been awarded to:

Professor Gunnar Jeschke

ESR Group

Laboratory of Physical Chemistry ETH Zürich Switzerland

The title of his lecture is

Measuring the Nanoworld

The lecture will take place in the City Suite at 20:00 on Tuesday 21st April. The wine at dinner on Tuesday evening and refreshments at the wine reception following the lecture are sponsored by Bruker UK. Previous winners of the Bruker Lectureship: 1986 MCR Symons 1995 H McConnell 2004 WL Hubbell 1987 K Möbius 1996 BM Hoffman 2005 K-P Dinse 1988 H Fischer 1997 KA McLauchlan 2006 YuD Tsvetkov 1989 JS Hyde 1998 JR Pilbrow 2007 D Goldfarb 1990 JH Freed 1999 J Schmidt 2008 E Groenen 1991 E de Boer 2000 D Gatteschi 1992 G Feher 2001 J Hütterman 1993 NM Atherton 2002 GR & SS Eaton (Joint) 1994 A Schweiger 2003 W Lubitz

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Next Meeting

The 43rd Annual International Meeting

of the

Electron Spin Resonance Spectroscopy Group

of the

Royal Society of Chemistry

will be held in

Cardiff

from 21st - 25th March 2010

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Upcoming Events

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International EPR/ESR Society

The Society is pleased to announce that the IES Awards for 2009 are:

Silver Medal for Chemistry: Takeji Takui. Silver Medal for Biology/Medicine: Gary Buettner.

Young Investigator's Award: Stefan Stoll.

For more information on the IES and how to join visit: http://www.ieprs.org/

or visit the epr newsletter site:

http://www.epr-newsletter.ethz.ch/

Copies of all Newsletters are available for download by members and a free public issue is available each year

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Programme

Mon 09:00 – 09:40

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Functional protein dynamics from Site Directed Spin Labeling

Wayne L. Hubbell University of California, Los Angeles

Jules Stein Eye Institute and Department of Chemistry and Biochemistry hubbellw@jsei.ucla.edu

By virtue of their small size and stabilization by weak interactions, protein molecules are inherently compressible and undergo substantial structural fluctuations. Evidence accumulated over the last decade shows that fluctuations on the ps to ms and longer time scales are functional, and determine in part the kinetics, specificity and affinity of ligand binding and protein-protein interactions, and provide a mechanism for allosteric coupling. To understand the molecular basis of protein function is then essential to have experimental tools available to detect motions on the above mentioned time scale. NMR relaxation methods have provided the most detailed description of dynamics for small proteins in solution, but face significant challenges in the study of more complex systems, i.e. transmembrane helical receptors in their native environment. Site Directed Spin Labeling (SDSL) and new strategies in EPR spectroscopy provide an alternative approach, not limited by the size and complexity of the system, with significant advantages of sensitivity and time scale. Earlier studies documented the ability of SDSL to map fast {ps-ns} backbone dynamics. Recent studies suggest that {µs-ms} conformational fluctuations can be detected in spin-labeled proteins through the use of osmotic and hydrostatic pressure perturbation, and exchange rates can be determined using pulse saturation recovery EPR spectroscopy.

Mon 09:45 – 10:00

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PELDOR studies of conformations of single- and double-stranded DNA with non-nucleotide inserts

S.A. Dzuba,* N.A. Kuznetsov,$ A.D. Milov,* , Yu.D. Tsvetkov,* O.S.

Fedorova$ *Institute of Chemical Kinetics and Combustion, Institutskaya 3, 630090,

Novosibirsk $Institute of Chemical Biology and Fundamental Medicine, Lavrentyev Ave.

8, 630090, Novosibirs) E-mail: dzuba@kinetics.nsc.ru

Specific structural conformational and energetic properties of DNA molecules containing damaged bases are believed to be the factors that enable repair enzymes to recognize these bases. The development of adequate experimental approaches to study these properties is desirable. In this work pulsed electron-electron double resonance (PELDOR) was applied to study conformations of double-spin-labeled single-stranded (ss) and double-stranded (ds) model DNA of 12-nucleotide length, containing in one strand the damaged (analog of apurinic/apyrimidinic site - tetrahydrofurane residue, F), or undamaged (guanine, G) base. Also, conformations were investigated of DNA which was kinked by introduction in the complementary strand the non-nucleotide inserts or additional nucleotides, forming a “loop” in the duplex. It was found that the distance distribution function n(r) for dsDNA is remarkably narrower, than that for ssDNA. Non-nucleotide inserts for the case of ssDNA only slightly influence n(r), while for dsDNA the insertion result in a noticeable bend of the molecule, with a bending angle of ~ 23o or ~ 27o, depending on the insert structure. The obtained results evidence that PELDOR may be successfully used as a “molecular ruler” for studying the influence of local damages on the DNA spatial structure and mechanical rigidity.

Mon 10:05 – 10:20

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Orientation selective DEER measurements in systems containing paramagnetic iron-sulfur clusters

J. Harmer*, JE Banham*, CR Timmel*, D Caprotti*, SG Bell*,I Forward*, LL Wong*,

F Mercuri¤, G. Jeschke$, Y Polyhach$ * Department of Chemistry, Centre for Advanced Electron Spin Resonance, ,

University of Oxford, UK ¤ University of Perugia, Department of Chemistry, Perugia, Italy

$ Laboratory of Physical Chemistry, Swiss Federal Institute of Technology, Zürich, Switzerland

jeffrey.harmer@chem.ox.ac.uk

Measurement of the magnetic dipole-dipole interaction between two unpaired electron spins provides a unique method to determine distance and orientation information in paramagnetic systems. DEER spectroscopy between two nitroxide radicals for instance is an established and reliable technique for distances in the range ca. 20

Mon 11:00 – 11:25

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Multifrequency Pulsed EPR Targeting Metal Cluster – Ligand Interactions

R. David Britt, Michelle M. Dicus, Gregory J.Yeagle, Troy A. Stich, & Stefan Stoll

Department of Chemistry, University of California, Davis 1 Shields Ave, Davis, CA, USA 95616

rdbritt@ucdavis.edu

We are using two mid-field/frequency pulsed EPR spectrometers to supplement our previous X-band EPR studies on metalloenzymes with paramagnetic metal clusters. One instrument is a lab-built high power 31 GHz pulsed spectrometer, and the other is a Bruker E580 with 34 GHz pulse capabilities. The pair of instruments allow us to reach the ESEEM “exact cancellation” limit for nitrogen ligands with stronger hyperfine couplings than those that fall in this regime in the more common X-band pulsed instruments. The mitoNEET protein has a 2Fe-2S cluster coordinated to a single histidine, providing an interesting motif midway between traditional all-cysteine liganded ferredoxins and the Rieske centers which have two histidine ligands. ESEEM at 31 and 34 GHz provides thorough characterization of the nitrogen couplings, and the results will be contrasted to parallel data on the Rieske center. Crystal structures of dinuclear manganese catalase cluster show two histidine ligands, but previous X-P band ESEEM revealed only one detectable nitrogen. New results with the midfield instruments reveal the second strongly coupled nitrogen. Finally, studies on the Mn cluster of Photosystem II show couplings to a single strongly coupled nitrogen that can be assigned to His332 of the D1 protein, consistent with the recent Barber/Iwata X-ray structure.

Mon 11:30 – 11:45

22

Engineering dioxygen sensitivity of hydrogenases : Mechanistic studies combining EPR, photochemistry and electrochemistry

B. Burlat*, F. Leroux*, E. Etienne*, S. Dementin*,

M. Rousset*, P. Bertrand*, C. Léger*, & B. Guigliarelli* *Bioénergétique et Ingénierie des Protéines, BIP-UPR9036, IMM-IFR88,

CNRS and Aix-Marseille University, 13402 Marseille, FRANCE guigliar@ifr88.cnrs-mrs.fr

Hydrogenases are enzymes that catalyze the reversible oxydation of H2 in the bioenergetic metabolism of many microorganisms. Potential application of hydrogenases concerns the biotechnological production of H2 but it is severely limited by the sensitivity of these enzymes to O2. A subclass of these enzymes contains a Ni-Fe dinuclear center as active site that is srongly buried in the protein and connected to the solvent through molecular tunnels. Both substrate (H2) and inhibitors (O2, CO) are considered to diffuse through these tunnels to reach the active site. Thus, rational strategies to improve the enzyme resistance were based on site-directed mutagenesis modifications of the tunnels to control the diffusion of inhibiteurs. In this work, we investigate the effect of mutations affecting the part of the tunnel located near the Ni-Fe active site. By using electrochemistry, we have shown that it is possible to measure the diffusion rates of gas within the enzyme (1). However, the results indicates that changes in O2 resistance of the enzyme cannot be accounted for by modifications of diffusion rates only (2). In order to analyze the molecular factors which trigger the hydrogenase reactivity with O2, the photochemical properties of the enzyme were studied by EPR spectroscopy. Upon illumination of the active enzyme at low temperature (< 100K), changes of the Ni-Fe EPR signal were observed which reveal light-induced transfer of hydrogenated species. The temperature dependence of the recombination process kinetics was studied by EPR. Our results show that, in addition to the restrictions of the tunnel size, the modification of O2 sensitivity induced by the mutations can be related to alteration of the H-bond network around the NiFe active site. References

1- Almeida MG, Silveira CM, Guigliarelli B, Bertrand P, Moura JJ, Moura I, Léger C. (2007) FEBS Lett, 581, 284-8.

2- Leroux F, Dementin S, Burlat B, Cournac L, Volbeda A, Champ S, Martin L, Guigliarelli B, Bertrand P, Fontecilla-Camps J, Rousset M, Léger C. (2008) Proc. Natl. Acad. Sci. U S A., 105, 11188-93.

Mon 11:50 – 12:05

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Multifrequency Electron Paramagnetic Study of the Radical Intermediates Generated by PpoA, a Novel CYP450 Fusion

Protein from Aspergillus nidulans

Alistair Fielding*, Florian Brodhun$, Ivo Feussner$ and Marina Bennati* *Max Planck Institute for Biophysical Chemistry, Am Faßberg 11, D-37077

Göttingen, Germany $Georg-August-University, Albrecht-von-Haller-Institute for Plant Science,

Department of Plant Biochemistry, Justus-von-Liebig-Weg 11, D-37077 Göttingen, Germany

afieldi@gwdg.de

PpoA is a fungal dioxygenase that produces hydroxylated fatty acids that are involved in the regulation of the life cycle of Aspergillus nidulans. This novel enzyme seems to harbor two functional heme domains consisting of an N-terminal peroxidase as well as a C-terminal CYP450 domain. It catalyzes the dioxygenation of linoleic acid to its 5,8-diol derivative via the formation of (8R)-hydroperoxy octadecadienoic acid [(8R)-HPODE]. We propose that the two reaction steps are catalyzed separately by the two different domains. In order to confirm this hypothesis we have expressed the protein in E. coli, purified it about 30-fold and used multifrequency EPR combined with freeze-quench experiments to study the radical intermediates possibly involved in the catalytic mechanism. Low temperature 9-GHz EPR spectra on samples prepared by dual mixing of the PpoA with either (8R)-HPODE or linoleic acid showed the presence on the millisecond time scale of a radical signal consisting of a doublet with a peak-to-trough of ~34 G. Simulation of the data at 95-GHz showed that the g-values were consistent with a hydrogen-bonded tyrosyl radical. Latter reaction times show the development of a second radical characterized by changes of line shape and a longer value of T1. The relative amounts of these signals vary between the two substrates. The origin of the radical signals and implications to the reaction mechanisms of this protein will be discussed.

Mon 12:10 – 12:25

24

Substrate radical intermediates in cyanobacterial bilin reductases

Stefan Stoll, Constantino P. Aznar, Wesley Sughrue, Shelley S. Martin, J. Clark Lagarias, R. David Britt

Department of Chemistry and Section of Molecular and Cellular Biology, University of California, Davis CA 95616

sstoll@ucdavis.edu Phycocyanobilin:ferredoxin oxidoreductase (PcyA) is a member of a group of bilin reductases that regioselectively reduce one or two double bonds of biliverdin IXα to produce linear tetrapyrrole chromophores that are used by organisms to capture light for photosynthesis and to detect light in phytochrome photoreceptors. In these bilin reductases, the reductions proceed via substrate-centred radical intermediates and involve successive steps of electron and proton transfers. The enzymes are highly unusual as they directly deliver the electrons from an extrinsic reductant to the substrate without the help of any cofactors such as flavins or iron-sulfur clusters.

We present recent results from high-field EPR, ENDOR and ESEEM of frozen solutions and single crystals of the trapped radical intermediate in wild-type and mutants of PcyA from Synechocystis sp. PCC6803 together with density functional theory calculations that help to identify the exact nature of the radical intermediates and thus enable us to clarify the reaction mechanism. [1] S. Stoll, A. Gunn, M. Brynda, W. Sughrue, A. C. Kohler, A. Ozarowski, A. J.

Fisher, J. C. Lagarias, R. D. Britt, J. Am. Chem. Soc. 2008, 131, 1986-1995. [2] T. Dammeyer, E. Hofmann, N. Frankenberg-Dinkel, J. Biol. Chem. 2008, 283, 27547-27554. [3] S.-L. Tu, H.-C. Chen, L.-W. Ku, J. Biol. Chem. 2008, 283, 27555-27564.

Mon 14:00 – 14:15 JEOL

25

Anisotropy in Molecular Magnetism – Magnetic Exchange Coupling of Octahedral Cobalt(II) Ions

A. B. Boeer,a F. Tuna,a G. Timco,a M. Baker,a A.-L. Barra,b R. Mole,c T. Unruh,c

E. J. L. McInnes,a D. Collisona and R. E. P. Winpennya aSchool of Chemistry, The University of Manchester, Manchester, M13 9PL, UK

bGHMFL, LCMI-CNRS, 38042 Grenoble, Cedex 9, France cZWE FRM-II, Lichtenbergstraße 1, Garching, 85747 München, Germany

angelika.boeer@postgrad.manchester.ac.uk Molecular nanomagnets have been studied intensively during the past 20 years and are still of growing interest towards the development of molecular magnetic storage devices, magnetic refrigeration and quantum information processing. A particularly important property to achieve a magnetic memory effect is a large axial magnetic anisotropy. One possibility to increase magnetic anisotropy is the use of magnetic ions with a unquenched orbital angular momentum such as octahedral cobalt(II). Spin-orbit coupling leads to a large effective splitting in zero-field, but a simplistic spin-only interpretation of magnetic exchange interaction is no longer sufficient. Very little is understood on the exchange coupling of cobalt(II) ions to date. A series of dimetallic cobalt(II) compounds, both ferro- and anti-ferromagnetically coupled, has been studied with various techniques on powder and single crystal samples. Detailed experiments include SQUID measurements, multifrequency EPR spectroscopy and inelastic neutron scattering and these provide information about the splitting in zero-field, and the effective "g-value" of the dimetallics. Such complementary data will be a major step towards the development of a theoretical model that describes the coupling within cobalt(II) oligomers. One experimental example is show below: The paramagnetic ground state of the cobalt (II) dimetallic [Co2(H2O)(O2CtBu)2(HO2CtBu)2(py)2] (structure in [1]) leads to unusually rich multifrequency EPR spectra. References

1. R. E. P. Winpenny et al, Chem. Eur. J. 2003, 9, 5142-5161.

0 1 2 3 4 5 6 7 8 9 10 11 12

magnetic field / T

345 GHz 285 GHz

230 GHz

190 GHz 94 GHz

34 GHz 9.4 GHz

Mon 14:20 – 14:35 JEOL

26

EPR and 55Mn-ENDOR Spectroscopy of the S2-State Multiline Signal of Photosystem II

Susanne Pudollek*, Sven Kessen, Christian Teutloff, Athina Zouni$, Jan Kern$ &

Robert Bittl *Freie Universität Berlin, Fachbereich Physik, Berlin

$Technische Universität Berlin, Max-Volmer-Laboratorium, Berlin Correspondence to robert.bittl@physik.fu-berlin.de

The oxygen-evolving complex (OEC) in Photosystem II (PSII) is the catalytic site for photosynthetic water oxidation. According to the present structural model based on X-ray crystallography [1], the core of the water-oxidising complex (WOC) consists of four Mn ions and one Ca ion. The bridging ligands between the metal ions and the protein ligands of the cluster are not unequivocally identified yet. Water oxidation occurs in five redox steps in the so-called S-state cycle (S0–4) [2] of the cluster. The foundation for a mechanistic understanding of the protein catalysed water-splitting and oxygen evolution at the OEC is the knowledge of the electronic structure of the Mn4Ca cluster and its ligand surrounding in the different S-states. The broad Multiline Signal (MLS) of the paramagnetic S2-state has already been extensively studied by various EPR techniques in different frequency bands and from different organisms [3–5]. Here, we present pulsed EPR spectroscopy at 94 GHz on PSII single crystals from Thermosynechococcus elongatus. 55Mn-ENDOR spectroscopy can directly provide information on the hyperfine couplings of the OEC and has so far mostly been performed in X- and Q-band for PSII prepared from spinach [6,7]. We present 55Mn-ENDOR experiments at 34 GHz on a frozen solution of PSII core complexes prepared from Th. elongatus. Species and preparation dependent differences in the 55Mn-ENDOR spectra are discussed. References

1. B Loll, J Kern, W Saenger, A Zouni & J Biesiadka; (2005) Nature 438, 1040–1044

2. B Kok, B Forbush & M McGloin; (1970) Photochem. Photobiol. 11, 457–475 3. DC Dismukes & Y Siderer; (1980) FEBS Letters 121, 78–80 4. C Teutloff, S Kessen, J Kern, A Zouni & R Bittl; (2006) FEBS Letters 580,

3605–3609 5. H Matsuoka, K Furukawa, T Kato, H Mino, JR Shen & A Kawamori; J. Phys.

Chem. B 110, 13242–13247 6. DW Randall, BE Sturgeon, JA Ball, GA Lorigan, MK Chan, MP Klein, WH

Armstrong & RD Britt; (1995) J. Am. Chem. Soc. 117, 11780–11789 7. LV Kulik, B Epel, W Lubitz & J Messinger; (2007) J. Am. Chem. Soc. 129,

13421–13435

Mon 14:40 – 14:55 JEOL

27

Exotica in pulsed dipolar spectroscopy – cobalt(II)-nitroxide distances and spatial distribution of nitroxides in membranes and

detergents

Bela E. Bode, Jörn Plackmeyer, Reza Dastvan, Thomas F. Prisner Institute of Physical and Theoretical Chemistry and Center for Biomolecular Magnetic Resonance, Goethe-University, Max-von-Laue-Straße 7, 60438

Frankfurt, Germany. bela@prisner.de

Pulsed electron-electron double resonance (PELDOR)1,2 methods have evolved to an important tool in measuring interspin distances in isolated nitroxide spin pairs.3,4 Pulse EPR distance measurements involving natural radical cofactors or transition metal ions have been performed, but are much less abundant in literature than distance measurements in bis-nitroxide labelled systems.4 In the context of our research on membrane transporters, their geometric arrangement and conformational flexibility in dependence of functional state, we were interested in the potential use of paramagnetic metal ions occupying the terminal his-tags used for protein isolation and purification. Therefore, we tested newly synthesized model systems consisting of a cobalt(II)-porphyrin covalently linked to a nitroxide for the feasibility of cobalt-nitroxide distance measurements. Deeply modulated PELDOR time-traces were obtained. The effects of conformational flexibility, spin-density distribution, orientation selection and g-anisotropy on the time domain data have been evaluated by explicit simulations. Furthermore, we investigated the effect of spatial distributions5,6 of spin labelled fatty acids in membrane vesicles and detergent. Due to the hydrophobic nature of these spinlabels they are expected to be inhomogeneously distributed through the sample. Distance measurements on these samples have gathered the spatial distribution functions and their comparison with analytic models. References

1. AD Milov, KM Salikhov & MD Shirov; (1981) Fiz. Tverd. Tela 23, 975; 2. AD Milov, AB Ponomarev & YD Tsvetkov; (1984) Chem. Phys. Lett. 110,

67. 3. G Jeschke & Y Polyhach; (2007) Phys. Chem. Chem. Phys. 9,1895. And

references therein. 4. O. Schiemann & TF Prisner; (2007) Quart. Rev.Biophys. 40, 1. And

references therein. 5. AD Milov, RI Samoilova, YD Tsvetkov, F Formaggio, C Toniolo & J Raap;

(2005) Appl. Magn. Reson. 29, 703. 6. AD Milov, DA Erilov, ES Salnikov, YD Tsvetkov, F Formaggio, C Toniolo &

J Raap; (2005) Phys. Chem. Chem. Phys. 7, 1794.

Mon 16:20 – 16:45

28

Doping of Perovskite Oxide Functional Materials: local structure determination using Electron Magnetic Resonance

D.J. Keeble,1 R.R. Garipov,1 J-.M. Spaeth,2 A. Peláiz-Barranco,3 V.V. Eremkin,4

and V. Smotrakov4 1Carnegie Laboratory of Physics, School of Engineering, Physics, and

Mathematics, University of Dundee, Dundee DD1 4HN, UK 2Fakultät Naturwissenschaften, Department Physik, Universität Paderborn, D-

33095 Paderborn, Germany 3Facultad de Física-Instituto de Ciencia y Tecnología de Materiales, Universidad

de La Habana, La Habana 10400, Cuba 4Institute of Physics, Rostov State University, Rostov on Don 344090, Russia

Recent years have seen a rapid growth in the development of functional materials based on the perovskite oxide, ABO3, structure. The ability to deposit epitaxial layers has led to the commercialisation of non-volatile Ferroelectric Random Access memory (FRAM) devices, mainly based on PbTiO3 related materials, and more recently to the observation of a two-dimensional electron gas at the interface between SrTiO3 and LaAlO3, which further stimulated interest in all-oxide electronic device structures. There has also been a resurgence of interest in bulk perovskite oxides, the development of piezoelectric engine fuel injectors, and the discovery of ultra-high strain single crystal piezoelectrics has been largely responsible for this. All these materials rely on the principles of point defect chemistry and physics to engineer the appropriate properties; impurity ion doping is widely used, but operates against a backdrop of a native and extrinsic distribution of defects. They all also suffer from a similar set of aging and fatigue mechanisms that are assumed to be driven by point defects. The aim of these studies has been to provide detailed local structure models of technologically relevant point defects, and ultimately use spectroscopic methods to investigate failure mechanisms. Here we demonstrate by pulsed ENDOR that Cu doping, i.e. acceptor ion doping, can lead to three different point defect configurations. We also discuss point defect structures determined by pulsed and conventional EPR methods in Fe3+ doped and rare-earth doped perovskite oxides.

Mon 16:50 – 17:05

29

EPR Analysis of Vanadium-doped Mesoporous Silica and Titania Catalysts

S. Zamani1, I. Caretti1, M. Chiesa2, V. Meynen3, E. Beyers3, A. Hanu3,4, P. Cool3, S.

Van Doorslaer1 1Department of Physics, University of Antwerp, Universiteitsplein 1, B-2610 Wilrijk-

Antwerp, Belgium 2Università di Torino and NIS Centre of Excellence, via P. Giuria 7, 10125 Torino,

Italy 3Department of Chemistry, Laboratory of Adsorption and Catalysis, University of

Antwerp, Universiteitsplein 1, B-2610 Wilrijk, Belgium 4Al. I. Cuza” University of Iasi, Department of Physical and Theoretical Chemistry

and Materials Chemistry, Bvd. Carol I, no 11, 700506, Romania Sabine.VanDoorslaer@ua.ac.be

Since 1992, a wide variety of mesoporous silica materials, such as MCM-41 and SBA-15, have gained growing interest, in order to overcome the limits in accessibility and applicability encountered for microporous zeolites. A lot of research has been done to activate these mesoporous materials with various transition metals to modulate the catalytic activity. In analogy, mesoporous titania materials are now being designed in order to combine the advantages of mesoporosity with the photocatalytic properties of titania materials. Here, we focus on the potential of different EPR techniques for the analysis of vanadyl-containing silica and titania materials. In a first part, we compare vanadium deposition in the pores of MCM-41 via the molecular designed dispersion (MDD) method with a facile, direct room-temperature synthesis targeted at vanadium incorporation into the MCM-41 framework. The difference between the two routes is monitored by studying the vanadyl-containing precursors of the final catalysts. It will be shown how important information about the mobility and incorporation mechanism of the vanadyl species in the mesoporous systems can be revealed by the use of X-band CW EPR and X- and W-band pulsed EPR spectroscopy. In a second part, these EPR techniques are applied to ‘mixed’ microporous-mesoporous materials, whereby vanadium silicalite-1 nanoparticles are deposited inside of the mesoporous channels of SBA-15. The effect of reduction under a H2 stream and subsequent addition of gasses, such as CO2 and NH3, is monitored and compared to the full-grown microporous silicalate. Finally, we show how similar pulsed EPR techniques and CW EPR in combination with light excitation can reveal information about the charge trapping and oxidation/reduction mechanisms of the vanadyl center and the titanate matrix in V-doped hydrogen titanate nanotubes.

Mon 17:10 – 17:25

30

An “EQR” spectrum: Electronic structure of rhenium tris(dithiolene) electron transfer series

Stephen Sproules and Eckhard Bill

Max-Planck-Institut für Bioanorganische Chemie, Stiftstrasse 34-36, D-45470 Mülheim an der Ruhr, Germany

sproules@mpi-muelheim.mpg.de The crystal structure of Re(pdt)3 (pdt = 1,2-diphenyl-1,2-dithiolate) was the first ever example of a coordination complex with trigonal prismatic geometry.1 As such, it was extensively studied by spectroscopic and theoretical techniques in an effort to identify the electronic reasons for its unusual geometry.2-4 Having an S = 1/2 ground state made it accessible to EPR spectroscopy, however, the spectrum is complicated and no attempts were initiated to explain these observations. We present here our X-band EPR study of the neutral Re(pdt)3 and the dianionic [Re(mnt)3]2-, (mnt = maleonitriledithiolate), and show that the complex spectral features result from a strong quadrupole interaction derived from a large valence contribution to the electric field gradient in conjugation with the sizable 183,185Re quadrupole moments. Spectral simulation for the neutral complex reveals the P-tensor is larger than the A-tensor, such that we classify this as an “electron quadrupole resonance (EQR)” spectrum. The result confirms the calculated electronic structure for the neutral complex as a low-spin Re(V) d2 with a π-radical ligand, while the two-electron reduced dianionic species is a low-spin Re(IV) d3 and three mnt2- ligands. References

1. R Eisenberg & JA Ibers; (1965) J. Am. Chem. Soc. 87, 3778-3779 2. EI Stiefel, R Eisenberg, RC Rosenberg & HB Gray; (1966) J. Am. Chem.

Soc. 88, 2956-2966 3. GN Schrauzer & VP Mayweg; (1966) J. Am. Chem. Soc. 88, 3235-3242 4. AH Al-Mowali & AL Porte; (1975) J. Chem. Soc., Dalton Trans. 250-252

Mon 17:30 – 17:45

31

CW and Pulsed ESR study on partially Zn (II) exchanged and partially hydrated Na-Zeolite A

C. Pal*, P.S. Wheatley#, H. EL. Mkami$, R. E. Morris#, O. Schiemann*

*Biomolecular Sciences, #EaStChem School of Chemistry,

$School of Physics and Astronomy, University of St Andrews, North Haugh, St Andrews, UK

cp70@st-andrews.ac.uk

Nitric oxide (NO) is a crucial biological agent. NO synthesised in the endothelial cells of blood vessels are found to mediate diverse functions like vasodilation, neurotransmission, inhibition of blood platelets.1 When produced in high concentrations it can contribute to inhibition of inflammation. Delivery of exogenous NO is therefore a promising way of treatment to various ailments. In order to achieve a target specific therapy new materials are required to be developed which can store significant quantities of NO and then deliver it at specific sites. Metal cation exchanged Zeolite frameworks have been manipulated for several uses because of their adsorbing property to several small molecules like CO, NO2, H2O or NO. NO adsorption on Na-A type of zeolites has been specifically investigated from catalysis point of view for NO decomposition and NO selective catalytic reduction.2 Partial exchange of Na-A with Zn (II) makes it more suitable to be used in physiological conditions due to less toxic Zn and increases its potential as a storage medium for NO. The NO adsorbed in the zeolite is paramagnetic and thus accessible to electron spin resonance (ESR) spectroscopy. In this study detailed continuous wave and pulsed hyperfine ESR measurements have been done on Zn exchange Na-A zeolite with and without NO to study the structure of the NO binding site to understand its storage capacity. References

1. PS Wheatley, AR Butler, MS Crane, S Fox, B Xiao, AG Rossi, IL Megson and RE Morris; (2006) J. Am. Chem. Soc. 128, 502-509.

2. A Pöppl, T Rudolf, P Manikandan and D Goldfarb; (2000) J. Am. Chem. Soc. 122, 10194-10200

Tues 09:00 – 09:40

32

Correlating the structure & function of proteins using EPR

Fraser MacMillan Henry Wellcome Unit for Biological EPR, School of Chemical Sciences and

Pharmacy, University of East Anglia, Norwich NR4 7TJ, UK, fraser.macmillan@uea.ac.uk

Electron paramagnetic resonance (EPR) spectroscopy is the ideal tool to study and characterise the functional role of intrinsic paramagnetic centres (eg organic radicals and transition metals) in electron transfer (ET) and other catalytic reactions of bioenergetic membrane protein complexes. Indeed we now have many specialised techniques at our disposal, which are used to identify and characterise in great detail such centres, eg high-field EPR, ENDOR and 2 dimensional pulse EPR methods like ESEEM, HYSCORE and more recently PELDOR (Pulsed ELectron DOuble Resonance). Further, spin labels specifically attached to such biomolecules (eg, via cysteines in proteins) allows EPR to now also access inherently diamagnetic systems and systematic use of pairs of any of the aforementioned spin-probes together with PELDOR allows structural information to be determined by studying their dipolar interactions. Our main goals are to correlate the structure and function of complex biomolecules including using spin labels and intrinsic paramagnetic centres to identify and characterise protein-protein interactions as a first step towards studying multi protein complexes or supercomplexes whose importance is only now starting to become apparent. The interfaces of most protein–protein complexes, however, are rather flat undulating surfaces with little regular topology thus it is unclear how a specific protein distinguishes an appropriate binding partner from several alternatives, which may contain very similar sequence and structure motifs. This discrimination is, however, of fundamental importance in many cell biological processes e.g. in signal transduction or within the immune system. In this talk I will show examples from our recent work on membrane proteins as well as on bacterial toxin-inhibitor protein complexes to illustrate the strengths of several of these techniques including some recent PELDOR work on multi-protein complexes.

Tues 09:45 – 10:00

33

DEER – probing nanosized self-assembled structures J. E. Lovett1,3, G. Jeschke2, S. M. Lea3, R. J. M. Abbott3, J. J. E. Caesar3, P. Roversi3, Y. Polyhach2, L. L. Wong4, D. Caprotti4, S. Bell4, I. Forward4, J. Harmer1, F. Mercuri4, S. D. Bell3, E. R. Barry3, A. Costa3, H. L. Anderson5, M. Hoffmann5, C. W. M. Kay6, A. Cnossen5, A. T. J. Shutter5, C. R. Timmel1

1Centre for Advanced ESR, Oxford, OX1 3QR 2ETH, Zürich, CH, CH-8093

3Sir William Dunn School of Pathology, Oxford, OX1 3RE 4Inorganic Chemistry Laboratory, Oxford, OX1 3QR

5Central Chemistry Laboratory, Oxford, OX1 3TA 6Biology Department, University College London, WC1E 6BT

janet.lovett@chem.ox.ac.uk This presentation will discuss four different self-assembled structures investigated by double electron electron resonance, DEER. Through these systems we will see how DEER is proving to be a useful tool for investigating structures which form in solution through weak, non-covalent bonds. 1. CD55 and factor B complex. Both proteins were spin-labelled and the structure of the overall complex mapped out by using the DEER distance restraints in docking programs. CD55 is attached to our cells and works to regulate our innate immune system so that we do not destroy our own cells but only actively target pathogens. Knowledge of the structure of immune system protein complexes will lead to effective targeted drug design as well as a deeper understanding of some hereditary diseases. 2. Ferredoxin and ferredoxin reductase complex in a Class I P450 electron transfer chain. One of the proteins contains a 2Fe2S cluster which can be made to have S = ½ while the other was selectively spin-labelled. The presence of the iron-sulfur cluster led to the requirement of analyzing the data in terms of selective excitation of orientations. 3. Architecture in a helicase. DEER was used to show the mechanism for the dynamic communication between secondary structure elements in the hexameric archaeal mini chromosome maintenance, MCM, helicase. The DNA replication machinery of archaea is particularly important since the principles are likely to be similar to the eukaryotic system and will therefore bring insight into tumour growth and cancer. 4. Non-biological nanowires. The series of wires ranged from containing one to four linked porphyrins with nitroxide spin labels at each end. The bending motion of the oligomers at two different temperatures was found by fitting the DEER signal. The technique was used to prove that the hypothesized self-assembled structures made from templating the wires, such as ladders and rings, were indeed formed.

Tues 10:05 – 10:20

34

Reciprocal transmembrane signaling in an ABC transporter

Mathias Grote1, Yevhen Polyhach2, Gunnar Jeschke2, Heinz-Jürgen Steinhoff3, Erwin Schneider1 and Enrica Bordignon2

1Institut für Biologie/Bakterienphysiologie, Humboldt Universität zu Berlin, Chausseestr. 117, D-10115 Berlin, Germany

2ETH Zurich, Laboratory for Physical Chemistry, Wolfgang-Pauli-Str. 10, 8093 Zurich, Switzerland

3Fachbereich Physik, Universität Osnabrück, Barbarastrasse 7, D-49076 Osnabrück, Germany

enrica.bordignon@phys.chem.ethz.ch In bacterial ABC importers the periplasmic anchoring of the substrate binding protein (receptor) is emerging as key determinant for the structural rearrangements in the cytoplasmically exposed ATP-binding cassette domains and in the transmembrane gates during the nucleotide cycle (1,2). Here the molecular mechanism of such signaling events was addressed by electron

paramagnetic resonance spectroscopy of spin-labeled ATP-binding cassette maltose transporter variants (MalFGK2-E) (3). Three functionally relevant conformations are found in the periplasmic MalF-P2 loop and concomitantly in the cytoplasmic ATP-binding cassette MalK2, strictly dependent on cytoplasmic nucleotide-binding and periplasmic docking of the receptor MalE to MalFG. In particular, a series of doubly spin-labeled mutants in the MalF-P2 domain and one triple mutant labeled at positions 205/252 in P2 and 83 in the Q-loop of MalK were assayed. The reciprocal communication unveiled here gives first insights into the stimulatory effect of MalE on the ATPase activity and it is suggested to be an important mechanistic feature of receptor-coupled ABC transporters in general (4,5).

References 1. Orelle, C., Ayvaz, T., Everly, R. M., Klug, C. S., and Davidson, A. L. (2008)

Proc. Natl. Acad. Sci. U. S. A. 105, 12837-12842 2. Goetz, B. A., Perozo, E., and Locher, K. P. (2009) FEBS Lett. 583, 266-270 3. Oldham, M. L., Khare, D., Quiocho, F. A., Davidson, A. L., and Chen, J. (2007)

Nature 450, 515-522 4. Locher, K. P. (2009) Philos. Trans. R. Soc. Lond. B Biol. Sci. 364, 239-245 5. Grote, M., Polyhach, Y., Jeschke, G., Steinhoff, H.-J., Schneider, E., and

Bordignon, E. (2009) submitted

Tues 11:00 – 11:25

35

Tuning redox activity by replacing the proximal heme ligand in cytochrome P450cam with selenocysteine

I. García-Rubio1, C. Aldag2, I. Gromov1, G. Jeschke1, and D. Hilvert2

Laboratory of Physical Chemistry1 and Laboratory of Organic Chemistry2, ETH-Zürich, 8093 Zurich (Switzerland) garciarubio@phys.chem.ethz.ch

Although the reaction mechanism of cytochromes P450 is still not fully understood, the remarkable monooxygenase activity of this large family of enzimes has been attributed to the heme containing active-site and the coordination of a cysteine thiolate to the heme cofactor. To probe this interaction in P450cam, cysteine has been replaced conservatively by selenocysteine. It turns out that the sulfur-to-selenium substitution subtly alters the structural and catalytic properties of the enzyme, providing a unique tool for probing P450 chemistry. Here we present the effects of the axial ligand substitution in the electronic properties of several stages in the enzymatic cycle. The two first states (the substrate-free and subatrate-bound resting state) contain ferric iron which is a useful probe for EPR studies. The effect of substrate binding on the spin density distribution was studied by determining the hyperfine interactions of the iron with the pirrole nitrogens using HYSCORE. The spin density distribution in the axial ligand was investigated by labelling the protein with 77Se and characterizing its hyperfine interaction with low frequency CW-EPR and ENDOR. We have also undertaken the study of other short-lived intermediates by fast freeze-quenching the reaction of the enzyme with peracids. In this stage of the enzymatic cycle the iron is EPR-silent but the effects of the sulfur-to-selenium substitution are observed indirectly by measuring the properties of the generated free-radical that interacts with the ferryl moiety.

Tues 11:30 – 11: 45

36

The electronic structure and protein interactions of YD• in

Photosystem II investigated by high-field ENDOR-spectroscopy 1Ch. Teutloff∗, 1S. Keßen, 2F. MacMillan, 3J. Kern, 3A. Zouni, 1R. Bittl

1Fachbereich Physik, Freie Universität Berlin, Arnimallee 14, D-14195 Berlin, Germany 2School of Chemical Sciences and Pharmacy, University of East

Anglia, Norwich, NR4 7TJ, United Kingdom 3Institut für Chemie, Technische Universität Berlin, Straße d. 17. Juni 135, D-10623 Berlin, Germany

*christian.teutloff@fu-berlin.de

The coupling of electron with proton transport is a common scheme in many enzymes and plays a central role in bioenergetics. The energy transduction for example in protein complexes in the respiratory chain and in the photosynthetic membrane is based mainly on this. Photosystem II in the latter contains a stable neutral tyrosine radical, YD• which is stabilized through deprotonation. This deprotonation step after radical formation can be monitored under certain conditions, thus forming an ideal model of a native radical in a protein undergoing proton-coupled electron transfer [1]. Here we present a detailed study of the stable YD•-radical in single crystals of PSII from Thermosynechococcus elongatus by EPR- and ENDOR-spectroscopy at 94 GHz. The higher sensitivity at high frequency enables the investigation of the small crystals with advanced pulsed methods, the site selection helps in the assignment of otherwise overlapping signals. This yields a complete picture about the spin density distribution in the radical via the proton hyperfine couplings and their assignment to the protons in the molecule. ENDOR on D2O-exchanged crystals yields exclusively the interactions of YD• with the protein. The analysis of the angular dependence of the 2H hyperfine couplings gives information about the association of the tyrosine with the environment. Furthermore EPR-investigations were undertaken on dark-adapted, Mn-depleted sample, were the YD• radical was formed by low-temperature illumination. It was shown by high-field EPR, that in this case the radical could not be stabilized via deprotonation [1]. ENDOR-spectroscopy can resolve hyperfine interactions to the OH-proton of the tyrosine. The potential rearrangement of the YD•-radical in the deprotonation step will be discussed.

[1] P. Faller, C. Goussias, A. W. Rutherford, S. Un, Proc. Nat. Acad. Sci 100 (2003)

Tues 11:50 – 12:05

37

Electron Spin Echo Study Of Molecular Motions of Spin Labeled Stearic Acids in a Model Phospholipid Bilayer

N.P.Isaev, S.A.Dzuba

Institute of Chemical Kinetics and Combustion, Institutskaya-3, 630090 Novosibirsk, Russia, and Novosibirsk State University, 630090, Pirogova-2,

Novosibirsk, Russia e-mail: isaev@gorodok.net

The dynamics of molecules in biological membranes are of the crucial importance for their functioning. Studies of molecular motions in biological membranes at cryogenic temperatures may allow detecting motions across low energetic barriers, which inevitably present, but cannot manifest itself at physiological temperatures. Fast stochastic librations, with correlation time at the nanosecond time scale, manifest itself in a two-pulse electron spin echo (ESE) experiment. Stimulated three-pulse ESE experiment is sensitive to motions of ultrasmall amplitude, ~ 0.1-1°, developing at the microsecond time scale. Investigation dynamics of DOXYL (4,4-dimethyl-oxazolidine-1-oxyl) labeled stearic acids in 1-Palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) shows that spin labels participate in stochastic orientational motions at temperatures down to ~120K for 5-DOXYL-stearic acid and down to ~80K for 16-DOXYL-stearic acid. The stimulated ESE decays were found to depend on the product of the two time delays of the pulse sequence. This fact may be described within a simple model of slow inertial rotations developing within this small range of angles with a rate of ~1 kHz. Both types of motion evidence the pronounced motional heterogeneity across the bilayer at cryogenic temperatures, with a remarkable increase of motion in the bilayer interior. The found low-temperature motions imply that hydrophobic parts of amphiphilic biomolecules may possess a noticeable mobility even at temperatures as low as ~100 K [1].

1. Nikolay P. Isaev and Sergei A. Dzuba; (2008) J. Phys. Chem. B 112, 13285–13291

Tues 12:10 -12:25

38

Using PELDOR to define and refine the domain orientation of modular proteins

R.Ward1, M. Zoltner1, L.Beer1, H.El Mkami2, I. Henderson3, T. Palmer1,

D.G.Norman1 1College of Life Sciences, University of Dundee, Dundee. 2School of Physics &

Astronomy, University of St Andrews, St Andrews. 3Division of Immunity & Infection, University of Birmingham

d.g.norman@dundee.ac.uk The outer membrane b-barrel trans-membrane proteins in mitochondria, chloroplasts and gram-negative bacteria, are folded into the membrane with the aid of polypeptide transport-associated (POTRA) domains. These domains occur, and probably function, as a tandem array situated on the periplasmic side of the outer membrane. Two crystal structure investigations and one NMR study have attempted to define the structure and articulation of the POTRA domains of the e.coli, prototypic Omp85 protein YaeT. Structural discrepancies both between the two related x-ray studies and the NMR study have left the question of domain-domain orientation unresolved. The ability of the PELDOR experiment to determine long-range distances between spin labels, in what are essentially solution conditions, is ideally suited to determining long-range domain-domain orientation. By comparing the distance distributions calculated from PELDOR data to synthetic distance distributions generated by using simple molecular dynamics on the NMR and crystal derived structures, we have been able to first, determine if any of the structures represent the solution structure accurately and secondly to quantify any differences. Using a simulated annealing refinement methodology in which the spin label is represented by an ensemble of positions, defining the available extent of spin label mobility, we have been able to demonstrate convergence to what we believe to be the true solution orientation of this two-domain protein. The determination of long-range orientation in modular proteins has application in a large number of biologically important systems. The approach we have demonstrated in this work will have important applications in many of these systems, extending and improving the results obtained with either x-ray crystallography or NMR.

Tues 20:00 – 21:00

39

Measuring the Nanoworld

G. Jeschke *ETH Zurich, Lab. Phys. Chem., Wolfgang-Pauli-Str. 10, 8093 Zurich, Switzerland

gunnar.jeschke@phys.chem.ethz.ch

Nearly 50 years have passed since Richard Feynman claimed that there was “Plenty of Room at the Bottom”.1 Chemists have approached this room from below by combining molecular building blocks to functional systems on nanometre length scales. The five decades since 1959 have also witnessed the rise of molecular biology. It was discovered that information in biomacromolecules is passed over length scales of a few nanometres, often by structural changes. Many important phenomena on these length scales are observable only in non-crystalline systems. They are described by soft matter physics, which was still unknown in 1959. The new nanoworld, envisaged by Feynman, has certainly become a field of great interest. This world is even more diverse than it was sketched in his talk. Yet, despite plenty of activity in this field, maps of the nanoworld are still full of white spots. The main reason for our continuing ignorance is a lack of precise tools for measurements on and between nanoobjects. Measurements by pulse EPR techniques can address nanometre distances in non-crystalline systems and are available since about 25 years.2 During the last decade they were combined with site-directed spin labelling techniques3 and access to distance distributions was obtained.4 Many incremental improvements in measurement technology and protocols have tremendously improved sensitivity and precision and have made such techniques widely available.5 We are now in a position to fill white spots in the map of the nanoworld. As a first example we look at flexibility of phenylene-ethynylene backbones, which are the most popular shape-persistent molecular building blocks. Extending on a preliminary study,6 data on the dependence of flexibility on temperature and structural details were obtained. As a second example we discuss the dimer structure of the Na+/H+ antiporter NhaA of Escherichia coli that was first obtained by EPR7 and is now supported by a new mutation and cryo transmission electron microscopy studies. By the third and last example, proline permease PutP of Escherichia coli, we show that sparse distance data allow to verify and extend a structural model for the helical bundle of a 54.3 kDa α-helical membrane protein with 13 transmembrane domains. References

1. R. P. Feynman, Engineering and Science, Caltech, 1960. see: http://www.zyvex.com/nanotech/feynman.html

2. A.D. Milov, A.B. Ponomarev, Yu.D. Tsvetkov: Chem. Phys. Lett. 110, 67 (1984)

3. W.L. Hubbell, D. S. Cafiso, C. Altenbach, Nature Struct. Biol. 7, 735 (2000) 4. G. Jeschke, A. Koch, U. Jonas, A. Godt, J. Magn. Reson. 155, 72 (2002) 5. G. Jeschke, Ye. Polyhach, Phys. Chem. Chem. Phys. 9, 1895 (2007) 6. A. Godt, M. Schulte, H. Zimmermann, G. Jeschke, Angew. Chem. Int. Ed.

45, 7560 (2006) 7. D. Hilger, Ye. Polyhach, E. Padan, H. Jung, G. Jeschke, Biophys. J. 93,

3675 (2007)

Weds 09:00 – 09:40

40

Quantum chemical calculations of EPR parameters as a tool to study metalloenzyme sites

Martin Kaupp

Institut für Anorganische Chemie, Universität Würzburg, Am Hubland, 97074 Würzburg, Germany, e-mail: kaupp@mail.uni-wuerzburg.de

Modern methods of density functional theory (DFT) have emerged over the past decade as an invaluable tool for explicit computation of EPR parameters (hyperfine and NQCC tensors, g-tensors and zero-field splittings), to complement and assist experimental studies in a wide range of fields.1 In my lecture, I will introduce some key aspects (e.g. exchange-correlation functional, basis set, proper consideration of spin-orbit and scalar relativistic effects) that need to be accounted for if one deals with transition-metal systems. These matters will be illustrated by example applications on metalloenzyme sites and models. These range from detailed evaluations of DFT methods to compute EPR parameters for Mo(V) and W(V) complexes2 to insights into the Cu hyperfine couplings in blue-copper enzyme sites.3 The most recent work concentrates on the application of broken-symmetry DFT approaches to multinuclear Mn sites: based on detailed validation studies on dinuclear Mn sites,4 the first explicit applications to the oxygen-evolving complex of photosystem II will be presented.4 References 1. Calculation of NMR and EPR Parameters. Theory and Applications (Eds. M.

Kaupp, M. Bühl, V. G. Malkin) Wiley-VCH, Weinheim 2004. 2. P. Hrobárik, O. L. Malkina, V. G. Malkin, M. Kaupp Chem. Phys. 2009, 356,

229. J. Fritscher, P. Hrobárik, M. Kaupp Inorg. Chem. 2007, 46, 8146. J. Fritscher, P. Hrobárik, M. Kaupp J. Phys. Chem. B 2007, 111, 4616.

3. C. Remenyi, R. Reviakine, M. Kaupp J. Phys. Chem. B 2007, 111, 8290. 4. S. Schinzel, M. Kaupp, submitted. 5. S. Schinzel, J. Schraut, M. Kaupp, submitted.

Weds 09:45 – 10:00

41

State space reduction and symmetry factorization in EPR and Spin Chemistry simulations

G.T.P. Charnock†, H.J. Hogben‡, P.J. Hore‡, I. Kuprov†

†Department of Chemistry, Durham University, South Road, Durham DH1 3LE ‡Department of Chemistry, Physical and Theoretical Chemistry Laboratory,

University of Oxford, South Parks Road, Oxford, OX1 3QZ

We report a number of theoretical developments aimed at improving the speed and efficiency of EPR and Spin Chemistry simulation algorithms. Specifically:

• Full permutation group factorization of the nuclear state subspace. We describe a semantic factorization procedure for the product operator basis and demonstrate that in a large number of EPR experiments and all Spin Chemistry experiments only the fully symmetric irreducible representation of the nuclear permutation group is ever populated. In particular, this means that "difficult" non-Abelian symmetry groups (Sn, Td, Dnh etc) can now be used in a straightforward way, since only the one-dimensional A1g irreducible representation is necessary. In the case of S4 symmetry in a six-spin radical pair, the initial 4096-dimensional Liouville state space is reduced to 79 states.

• Coherence order screening in isotropic singlet-source MARY (magne-

tically altered reaction yield) simulations. All isotropic photo-generated MARY systems have SO(2) symmetry, meaning that spin dynamics is confined to the fairly small subspace with totalZ 0L = .

• The location and properties of large unpopulated and weakly populated

subspaces in the state spaces encountered in EPR and Spin Chemistry. The subspaces in question arise from the spin system interaction structure (which is never all-to-all) and the rarely exploited non-geometric and continuous symmetries that exist in EPR spin systems.

The theoretical results listed above are implemented in the forthcoming release of the open-source Spinach libraries, which aim to automate state space reduction and provide easy-to-use programming tools for the generation of low-dimensional representations in the simulations of large spin systems.

Weds 10:05 – 10:20

42

Revisiting the McConnell relationship: why C1?

Dimitri Svistunenko, Garth Jones Department of Biological Sciences, University of Essex, Wivenhoe Park,

Colchester, EssexCO4 3SQ. svist@essex.ac.uk The EPR spectrum lineshape of tyrosyl radicals depends strongly on the hyperfine interactions between the unpaired electron and the methylene protons Hβ1 and Hβ2. The isotropic part of the hyperfine splitting constant for the protons Aβjiso (j = 1, 2) can be estimated from the semi-empirical McConnell relationships

Aβj

iso = ρ

C1(B’ + B’’cos

2θ

j) (1)

where ρc1 is the spin density on atom C1, θj is the angle for the j-th proton (this angle is indicated on the Figure for proton Hβ1) and B’ and B‘’ are constants. Density functional theory (DFT) can be used to calculate both the spin densities on different atoms of a radical and the hyperfine interaction constants. The McConnell relationship can further be used to calculate the spin density on atom C1 from the hyperfine interaction constants. One would expect that thus calculated ‘McConnell spin density’ on C1 correlates with the spin density on this atom directly calculated by DFT (the Mulliken spin density). We have performed the DFT calculation for tyrosyl radicals of four different enzymes and for a hydrogen bonded tyrosyl radical model, at different levels of theory. Surprisingly, calculated McConnell spin density on C1 does not correlate with the Mulliken spin density on this atom. Instead a clear negative correlation was found with the Mulliken spin density on the phenoxyl oxygen (Figure). We therefore suggest that the McConnell relationship can be modified as follows:

Aβj

iso = (1-ρ

O)(C’ + C’’cos

2θ

i) (i=1,2) (2)

This equation uses the spin density on the phenoxyl oxygen rather than on C1. The (1-ρ

O) factor represents approximately the total spin density on the ring

excluding the density on the phenoxyl oxygen. Thus, optimization of parameters C’ and C’’ in Equation (2) for different tyrosyl radicals would yield more consistent values (less dispersed) than optimization of parameters B’ and B’’ in Equation (1). The reasons why ρ

O would better describe the A values than ρ

C1 will be discussed.

Weds 11:00 – 11:25

43

1H and 13C Dynamic Nuclear Polarisation with a Two-Field (0.35/14 T) Shuttle Spectrometer

M. Bennatia , M.T. Türkea, M. Reesea, I. Tkacha, C. Griesingera, G. Parigib, C. Luchinatb, P. Höferc, A. Tavernierc, T. Marquardsenc, F.

Engelkec aMax Planck Institute for Biophysical Chemistry, Göttingen, bCERM, University of

Florence, Sesto Fiorentino, Italy, cBruker Biospin, EPR Division, Rheinstetten, Germany

bennati@mpibpc.mpg.de

Dynamic nuclear polarization (DNP) permits to increase the NMR signal of nuclei by pumping the electronic spin transitions of paramagnetic centers nearby1. This method is emerging as a powerful tool to increase the inherent sensitivity of NMR in structural biology aiming at detection of macromolecules. In liquid solutions, DNP is governed by the Overhauser effect, which depends on various physical parameters but overall looses efficiency with increasing magnetic fields. In aqueous solution, additional technical issues associated with the penetration of microwaves in water and heating effects aggravate the performance of the experiment. We have recently reported large 1H-DNP enhancements (> 100) on water solutions containing the nitroxide spin label 4-hydroxy-amino-TEMPO (TEMPOL) by using state-of-the-art microwave technology at 9 GHz electron pumping frequencies2,3. To examine the feasibility of low-field (9 GHz/0.35T) DNP in high resolution NMR, we have constructed the prototype of a two-field shuttle DNP spectrometer that polarizes nuclei at 9 GHz/0.35 Tesla and detects the sample polarization at 14 Tesla4. We now report our first 1H and 13C DNP results with this spectrometer. The sign of the enhancements gives insight into the acting DNP mechanism. The results provide a proof of principle for the feasibility of a shuttle DNP experiment and open up perspectives for the application potential of this method in solution NMR. References

1. Abragam, A. Principles of Nuclear Magnetism; University Press: Oxford, 1961.

2. Höfer, P.; Parigi, C.; Luchinat, C.; Carl, P.; Guthausen, G.; Reese, M.; Carlomagno, T.; Griesinger, C.; Bennati, M. J. Am. Chem. Soc. 2008, 130, 3254-3255.

3. Höfer, P.; Carl, P.; Guthausen, G.; Prisner, T.; Reese, M.; Carlomagno, T.; Griesinger, C.; Bennati, M., Appl. Magn. Reson. 2008, 34, 393-398.

4. Reese, M.; Lennarrtz, D.; Marquardsen, T.; Höfer, P.; Tavernier, A.; Carl, P.; Schippmann, T.; Bennati, M.; Carlomagno, T.; Engelke, F.; Griesinger, C. Appl. Magn. Reson. 2008, 34, 301-311.

Weds 11:30 – 11:45

44

Very high bandwidth, orientation selective, DEER spectroscopy at 94GHz

G.M.Smith1, P.A.S.Cruickshank1, O.Schiemann2, D.R.Bolton1, D.A.Robertson1,

R.I.Hunter1, H.El Mkami1 *1School of Physics and Astronomy, University of St Andrews, Scotland

2 School of Biology, University of St Andrews, Scotland E-mail: gms@st-and.ac.uk

Pulsed Double Electron-Electron Resonance (DEER) spectroscopy in combination with site directed spin labeling, is now well known as a powerful technique for accurate long-range distance measurements in biomolecules, up to 5 nm and beyond. Today, the vast majority of these measurements are run at 10 GHz using commercial instrumentation, as this setup currently offers the highest stability and sensitivity. More recently there has been interest in moving to sufficiently high magnetic fields to allow the g-anisotropy of nitroxides to become clearly resolved, in order to be sensitive to the relative orientation of the nitroxide pairs. Proof of principle experiments have recently been carried out at 180 GHz1 and 95 GHz2, which have clearly indicated the potential of this methodology. However, these experiments have also suffered from low sensitivity (relative to X-band) and low excitation and detection bandwidths due to the relatively low source power available, which in turn have required the use of high Q cavities. At St Andrews, a high power kW pulse system at 94GHz has been developed with extremely low deadtime and virtually no standing waves. This system can deliver effective 5 ns π/2 pulses to non-resonant sample holders (of high volume), at any desired frequency over 1GHz instantaneous bandwidths. This allows near optimal pulses to be delivered for DEER experiments at any frequency across the full 500 MHz spectral width of a nitroxide spectrum at 94 GHz. This not only offers high sensitivity, but allows a full set of measurements that permit all relative nitroxide pair orientations to be correlated separately and gives a set of measurements, which now becomes very sensitive to small angular shifts in the relative angular positions of the nitroxide pairs. This methodology holds great promise in being able to characterise small conformational changes in biomolecules even in the presence of broad distance distributions or even where the nitroxides have a large inherent degree of flexibility in their relative orientation. In principle, the technique should become extremely sensitive to relative changes in orientation in cases where the nitroxide positions are fixed relative to each other. In this paper, we will outline the general experimental methodology and analysis and give recent experimental results that indicate the potential of the technique. References

2. V.P.Denysenkov, T.F.Prisner, J.Stubbe, M.Bennati, PNAS, Vol. 103, No.36, September, 2006, pp.13386-13390

3. Ye. Polyhach, A.Godt, C.Bauer, G. Jeschke, Journal of Magnetic Resonance, Vol.185, Issue 1, March 2007, pp.118-129

Weds 11:50 – 12:05

45

ENDOR in field-frequency space: orientation, species and quantum state selection

H. Vrielinck,* F. Loncke, A. Tarpan,$ H. De Cooman,$ D. Zverev & F. Callens

Ghent University – Dept. of Solid State Sciences, Krijgslaan 281-S1, B9000 Gent, Belgium,

*Postdoctoral Fellow and $Research assistant of the Flemish Research Foundation (FWO-Vlaanderen)

Henk.Vrielinck@UGent.be

Due to its specific detection method, via saturation of the EPR spectrum at a certain magnetic field position, ENDOR measurements are highly selective. Already in the late sixties Rist and Hyde recognized the possibilities of obtaining angular dependent information from powder specimens by recording the field-dependence of the ENDOR spectrum, by grace of the orientation selection principle.1 However, also for single crystal samples it makes sense to record ENDOR spectra in the two-dimensional field-frequency space (FF-ENDOR). Next to orientation selectivity – when several symmetry-related orientations of the same paramagnetic species are simultaneously detected in the EPR spectrum – such measurements feature species selectivity and, for systems with S>½ and/or I>½, also quantum state selectivity. The former facilitates the interpretation of multi- composite EPR spectra, as illustrated in the figure below. Quantum state selectivity offers possibilities of determining the relative signs of spin Hamiltonian parameters, e.g. zero-field splitting and (super)hyperfine, or (super)hyperfine and quadrupole principal values. All these effects will be illustrated through recent examples in our research of radiation-induced radicals in sugars and high-spin transition metal or rare-earth doped halide crystals.

Q-band (33.97 GHz) Field-Frequency ENDOR spectrum of sucrose immediately after X-ray irradiation at room temperature. Transitions

assigned to the stable (T1-3) 2 and unstable (U1-3) radicals are labelled. References

4. GH Rist & JS Hyde; (1970) J. Chem. Phys. 52, 4633-4643 5. H De Cooman, E Pauwels, H Vrielinck, A Dimitrova, N Yordanov, E

Sagstuen, M Waroquier & F Callens, Spectrochim. Acta A 69, 1372-1383

Weds 12:10 – 12:25

46

Development of high field OMRI scanner for imaging in vivo redox status in mouse

Hideo Utsumi, Kazuhiro Ichikawa and Tatsuya Naganuma Faculty of Pharmaceutical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka, 812-8582 JAPAN

utsumi@pch.phar.kyushu-u.ac.jp

In vivo redox status are repeatedly involved in oxidative diseases. The redox imaging technique is thus important to diagnose redox-induced diseases and to access cure effects of pharmaceutical drugs. OMRI is a new technique for imaging free radicals / redox status in animals based on the Overhauser effect1-3 and enables simultaneous redox imaging in animal4. The detection process is the same as that in MRI and the high resolution of MRI can be utilized in free radical imaging. The large difference of gyromagnetic ratio between electron and proton sp