nucleic acids andmolecular biology 14978-3-642-18851-0/1.pdfnucleic acid interactions, e.g., how...

Series EditorH. J. Gross

Nucleic Acids and Molecular Biology

14

Springer-Verlag Berlin Heide1berg GmbH

Alfred M. Pingoud (Ed.)

RestrictionEndonucleases

With 100 Figures, 27 of Them in Color

Springer

Professor Dr. ALFRED M. PINGOUD

Institute for Biochemistry Justus-Liebig-University Heinrich-Buff-Ring 58 35392 Giessen Germany

ISSN 0933-1891

ISBN 978-3-642-62324-0

Library of Congress Cataloging-in-Publication Data

Restriction endonudeases / Alfred M. Pingoud. p. cm. -- (Nucleic acids and molecular biology)

Includes bibliographical references and index. ISBN 978-3-642-62324-0 ISBN 978-3-642-18851-0 (eBook) DOI 10.1007/978-3-642-18851-0

1. Restriction enzymes, DNA. 2. Endonucleases. 3. Gene amplification. 1. Pingoud,A. (Alfred) Il. Series.

QP609.R44R4732003 572'.785--dc22

2003064952

This work is subject to copyright. AII rights are reserved, whether the whole or part of the material is concerned, specifically the rights of translation, reprinting, reuse of illustrations, recitation, broadcasting, reproduction on microfilm Of in any other way, and storage in data banks. Duplication of this publication or parts thereof is permitted only under the provisions of the German Copyright Law of September 9, 1965, in its current version, and permissions for use must always be obtained from Springer-Verlag. Violations are liable for prosecution under the German Copyright Law.

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Preface

Approximately 50 years ago the phenomenon of restriction was firstdescribed. This led later to the discovery of restriction (and modification)enzymes by Werner Arber, Daniel Nathans and Hamilton Smith who receivedthe Nobel Prize in Physiology or Medicine 1978"for the discovery of restriction enzymes and their application to problems of molecular genetics" . In retrospect, it is clear that the impact of this discovery was much greater: therewould be no recombinant DNA revolution and no gene technology withoutrestriction endonucleases. However,beyond their tremendous importance astools for the analysis and recombination of DNA, restriction enzymes haveprovided outstanding model systems to study many aspects of proteinnucleic acid interactions, e.g., how proteins find their target sites within agreat excess of non-specific sites, how short DNA sequences are recognizedwith such extreme accuracy, and how recognition is so efficiently coupled tocatalysis.

Restriction enzymes are usually part of restriction-modification (R-M)systems that serve to protect bacteria (Prokarya as well as Archaea) againstphage infections and uptake of foreign DNA. There are three main types (I,II and III) of restriction enzymes, which differ in subunit composition,cofactor requirement and mode of action. The best-studied (and most simple) ones are the Type II restriction endonucleases of which more than 3500are known . It was not recognized initially that restriction endonucleases areevolutionarily related, because - with few exceptions - they have little if anysequence homology. Only after the first crystal structures were determineddid it become clear that most Type II restriction endonucleases are relatedin evolution and that they share a similar active site with Type I and IIIrestriction endonucleases. The diversity of restriction enzymes makes themfascinating objects for studying the evolution of a family of enzymes with acommon basic function - highly specific DNA recognition and cleavage.Surprisingly, with probably only very few exceptions, they do not seem to berelated to another family of site-specific endonucleases, the homing endonucleases.

VI Preface

In the last few years, considerable progress has been made regarding themechanism of action of restriction enzymes . Still, many open questionsremain, even very basic ones. For example, there is not yet a consensus on themechanism of target site location or the details of the mechanism of DNAcleavage,and only little is known about the conformational changes followinginitial DNA binding by restriction endonucleases and leading to the activation of their catalytic centers. Major challenges remain, the most ambitiousone being to use the knowledge that we have regarding structure-functionrelationships of these enzymes to engineer variants with new specificities.

I am very pleased that so many of the top scientists in the field could beconvinced to provide chapters and I am grateful to them for taking the time todo so. The result of their efforts is a book that represents the most detailedaccount of restriction endonucleases currently available.

Giessen, September 2003 AlfredPingoud

Contents

Survey and SummaryA Nomenclature for Restriction Enzymes,DNAMethyltransferases, Homing Endonucleases and Their Genes 1

R.J. ROBERTS, M. BELFORT, T.BESTOR, A.S.BHAGWAT, T.A.BICKLE,J.BITINAITE, R.M. BLUMENTHAL, S.K.DEGTYAREV, D.T.F. DRYDEN,K. DYBVIG, K.FIRMAN, E.S.GROMOVA, R.I. GUMPORT, S.E. HALFORD,S. HATTMAN, J.HEITMAN, D.P. HORNBY, A. JANULAITIS, A. JELTSCH,J.JOSEPHSEN, A. KISS, T.R.KLAENHAMMER, I. KOBAYASHI, H. KONG,D.H. KRUGER, S. LACKS, M.G.MARINUS, M. MIYAHARA, R.D.MORGAN,N.E. MURRAY, V. NAGARAJA, A. PIEKAROWICZ, A. PINGOUD, E. RALEIGH,D.N.RAO, N. REICH, V.E. REPIN, E.D.SELKER, P.-c. SHAW, D.C.STEIN,B.L. STODDARD, W. SZYBALSKI, T.A. TRAUTNER, J.L. VAN ETTEN,J.M.B. VITOR, G.G. WILSON, S.-Y. Xu

1233.13.23.33.43.53.63.73.83.93.103.113.123.13

Introduction . . . . . . .General Rules .Details of Types and SubtypesTypes I, II, III and IVType I ..Type II .Type lIPType IIAType lIBType IICType lIEType IIFType IIGType IIHType 11MType lIS

3366678

1010111111

11121212

VIII

3.14 Type IIT .3.15 Nicking Enzymes .3.16 Control Proteins .3.17 Type III .3.18 Type IV .3.19 Hypothetical Enzymes3.20 Homing Endonucleases3.21 Adherence to These Conventions and UpdatesReferences . . . . . . . . . . . . . . . . . . . . . . . . . .

Restriction-Modification Systems as Minimal Forms of Life

I. KOBAYASHI

Contents

121213141415151616

19

122.12.2

2.32.42.4.12.4.22.4.32.4.42.4.5

2.53

3.13.2

3.33.44

4.14.24.2.1

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . .Genomics and Mobility of Restriction-Modification SystemsGenomics .Horizontal Gene Transfer Inferredfrom Evolutionary Analyses .Presence on Mobile Genetic Elements . . . . .Genomic Contexts and Genome ComparisonInsertion into an Operon-Like Gene Cluster .Insertion with Long Target Duplication . .Substitution Adjacent to a Large Inversion .Apparent Transposition . . . . . . . . . . . . . . . . .Linkage of a Restriction-Modification Gene Complexwith Another Restriction-Modification Gene Complexor a Cell Death-Related Gene .Defective Restriction-Modification Gene ComplexesAttack on the Host Genome and the SelfishGene Hypothesis .Post-Segregational Host Cell Killing .Comparison with other Post-SegregationalCell Killing Systems . . . . . . . . . . . . .Selfish Gene Hypothesis .Genomics as Explained by the Selfish Gene HypothesisGene Regulation in the Life Cycleof Restriction-Modification SystemsGene Organization .Gene Regulation . . . . . . . . . . . .Restriction Gene Downstream of Modification Gene

192222

23232326272727

2727

2828

293233

34363737

Contents IX

9

5

6

6.1

37373838

545556

495152

53

39

43

45484849

42

44

42

40

43

40

Restriction Gene Upstream of Modification GeneModification Enzyme as a RegulatorRegulatory Proteins . . . . . . . . . . . . .Type I Restriction-Modification Systems .Restriction-Modification Gene ComplexesMay Be Able to Multiply ThemselvesIntra-Genomic Competition InvolvingRestriction-Modification Gene Complexes .Two Restriction-Modification Systems withthe Same Recognition Sequence Can Blockthe Post-Segregational Killing Potential of Each OtherSolitary Methyltransferases Can Attenuatethe Post-Segregational Killing Activityof Restriction-Modification Systems .Resident Restriction-Modification Systems Can Abortthe Establishment of a Similar IncomingRestriction-Modification System .Suicidal Defense Against Restriction-ModificationGene Complexes . . . . . . . . . . . . . . . . . . . . . . . . .Defense Against Invaders byRestriction-Modification Systems .Genome Dynamics and Genome Co-Evolutionwith Restriction-Modification Gene ComplexesSome Restriction-Modification Gene Complexesand Restriction Sites Are Eliminated from the GenomeMutagenesis and Anti-Mutagenesis .End Joining .Homologous Recombination by BacteriophagesCellular Homologous Recombination in Conflictand Collaboration with Restriction-ModificationGene Complexes . . . . . . . . . . . . . . . . . . .Selfish Genome Rearrangement Model .Towards Natural Classification of Restriction EnzymesApplication of the Behavior of Restriction-ModificationGene Complexes as Selfish Elements .A Hypothesis on the Attack by Restriction-ModificationGene Complexes on the Chromosomes

10 ConclusionsReferences . . . . . . . . . . . . . . . . . . . . . .

5.5

5.4

5.2

5.3

5.1

6.678

4.2.24.2.34.2.44.2.54.3

6.26.36.46.5

x

Molecular Phylogenetics of Restriction Endonucleases

I.M.BUJNICKI

Contents

63

1 Discovery and Classification of Restriction Enzymes 632 Genomic Context of R-MSystems . . . . . . . . 643 Historical Perspective of Comparative Analyses

of Restriction Enzymes: Are They Productsof Divergent or Convergent Evolution? ... . . 66

4 Crystallography of Type II REases: Explorationof the "Midnight Zone of Homology" . . . . . . 66

5 Homology Between Restriction Endonucleasesand Other Enzymes Acting on Nucleic Acids . . 70

6 Non-Homologous Active Sites in Homologous Structures. 717 Cladistic Analysis of the PD-(D/E)xK Superfamily . . . . . 748 Identification of PD-(D/E)xK Domains in Other Nucleases

and Prediction of Their Position on the Phylogenetic Tree 799 In the End, Convergence Wins: Sequence Analyses Reveal

Type II Enzymes Unrelated to the PD-(D/E)XK Superfamily 8310 Evolutionary Trajectories of Restriction Enzymes:

Relationships to Other Polyphyletic Groups of Nucleases 84References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 87

Sliding or Hopping? How Restriction EnzymesFind Their Wayon DNA .

A. IELTscH, C. URBANKE

95

12

2.12.22.33

44.14.2

4.3

Introduction . . . . . . . . . . . . . . . . . . . .Mechanisms of Facilitated Target Site Locationby Proteins on DNA .Sliding .Hopping .Intersegment TransferCritical Factors Determining the Efficiencyof Target Site Location by Sliding and Hopping ProcessesSliding or "Hopping" - A Survey of Experimental DataStructures of Restriction Endonucleases .Accurate Scanning of the DNAfor the Presenceof Target Sites .Length Dependence of Linear Diffusion

95

96979899

100102102

102103

Contents XI

4.4 Processivity of DNA Cleavage 1044.5 DNA Cleavage by a Covalently Closed EcoRVVariant . 1064.6 Cleavage of Topological Connected Plasmid Molecules 1075 Conclusions 107References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 108

The Type I and III Restriction Endonucleases: StructuralElements in Molecular Motors that Process DNA . ... .

S.E. MCCLELLAND, M.D. SZCZELKUN

III

1 Energy-Dependent DNA Processing . . . . . . . . . 1112 Motor Enzyme Architecture of the ATP-Dependent

Restriction Endonucleases 1132.1 Motor Enzyme Motifs in the Type I and III

Restriction Endonucleases 1132.1.1 Gross Organisation of the Type I HsdR Subunits 1172.1.2 Gross Organisation of the Type III Res Subunits 1192.1.3 Core Helicase Motifs in ATP Binding and Catalysis 1192.1.4 The "Q-Tip Helix" - A New Helicase Motif? . . . . . 1202.1.5 The DNA Binding Motifs - Family-Specific Deviations 1212.2 A Motor Enzyme Fold in the Type I and III

Restriction Endonucleases 1232.3 Macromolecular Assembly of the Type I and III Restriction

Endonucleases . . . . . . . . . . . . . . . . . . . . 1253 Future Directions 1283.1 Coupling Chemical Energy to Mechanical Motion . . . 1283.2 Tools for Nanotechnology Rather Than Biotechnology? 131References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 132

The Integration of Recognition and Cleavage: X-Ray Structuresof Pre-Transition State Complex, Post-Reactive Complexand the DNA-Free Endonuclease 137

A. GRIGORESCU, M. HORVATH, P.A. WILKOSZ, K. CHANDRASEKHAR,J.M. ROSENBERG

12

Introduction . . . . . . . . . . . .The Pre-Transition State Complex

137139

XII Contents

2.1 General Features 1392.2 DNA Numbering Scheme 1402.3 Secondary Structure 1402.4 The EcoRI Kink 1422.5 Recognition Overview 1422.6 Sequence-Specific Hydrogen Bonds 1432.7 Sequence-Specific Interactions Via Bound Water . . 1472.8 Sequence-Specific Van der Waals Interactions .... 1472.9 Redundancy of Direct Sequence-Specific Interactions 1482.10 Bound Solvent . . . . . . . . . . . . . . . . . . 1492.11 "Buried" DNAPhosphate Groups 1492.12 Solvent-Mediated Contacts to the DNABases

Flanking the Recognition Site 1512.13 DNAMinor Groove ... . . . . . . . . . . . . 1522.14 Contrasting the 1.85and 2.7 APre-Transition

State Complexes . . . . . . . . . . . . . . . . . . . . . . . . 1533 The Post-Reactive Complex and the Cleavage Mechanism 1543.1 EcoRI Endonuclease Crystal Packing . 1543.2 Catalytic Site . . . . . . . . . . . . . . . . . . . . . . 1553.3 The Proposed CleavageMechanism . . . . . . . . . 1583.4 The Integration of EcoRI-Catalyzed DNACleavage

with Substrate Recognition . . . . . . . 1623.5 Chimeric PSheets . . . . . . . . . . . . 1644 Apo-Eco RI Endonuclease (The Protein

in the Absence of DNA) . . . . . . . . . 1664.1 Three Polypeptide Chains in the Asymmetric Unit 1664.2 Disordered "Arm" Regions 1674.3 Relation to Crystallographic Packing 1674.4 Order-Disorder Transition Associated with DNABinding 1695 Summary and Conclusions 172References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 173

Structure and Function of EcoRV Endonuclease

EK.WINKLER, A.E. PROTA

179

122.1

2.1.1

Introduction . . . . . . . . . . . . . . . . . . . . . .Structural Characteristics of EcoRV EndonucleaseDNA-Protein Interactions in Specificand Nonspecific Complexes ..The Specific DNABinding Mode . . .

179180

184185

Contents XIII

2.1.2 The Nonspecific DNABinding Mode 1882.2 The Structure of the ActiveSite 1892.2.1 The Structure in the Absence of Divalent Cations 1892.2.2 The Location of Bound Divalent Metal Ions 1912.2.3 The Structure of the Enzyme Product Complex . 1913 Biochemical Characteristics of EcoRV Endonuclease

and Structure-Function Relationships 1923.1 Thermodynamics and Energetics of DNABinding 1933.1.1 Specific and Nonspecific Binding to DNA . . . . . . . . 1933.1.2 The Energetics of Direct and Indirect Readout Interactions 1963.2 Chemistry and Kinetics of DNACleavage . . . 1993.2.1 General Aspects of Phosphodiester Hydrolysis 1993.2.2 Steady State and Rapid Reaction Kinetics . . . 1993.3 Mechanistic Hypotheses 2013.3.1 The Role of the Conserved, Catalytic-Motif Residues 2023.3.2 The Role of Metal Ions and the Need

of Conformational Changes in Different Mechanisms 2023.3.3 Catalytically Active and Inactive Conformational States 2043.3.4 The Three Metal Ion Mechanism 2084 Conclusions 209References . . . . . . . . . . . . . . . . . . . 210

Twoof a Kind: BamHI and BglII

E. SCHEURING VANAMEE, H.VIADIU, C.M. LUKACS, A.K. AGGARWAL

215

1 Introduction . . . . . . . . . . 2152 BamHI Endonuclease . . . . . 2162.1 The Structure of Free BamHI 2162.2 BamHI Nonspecific Complex 2172.3 BamHI Specific Complex . . . 2192.4 The Pre-Reactive Complex of BamHI 2242.5 The Structure of the BamHI Post-Reactive Complex. 2253 BglII Endonuclease 2273.1 The Structure of Free BglII . . . . . . . . . . . . . . . 2273.2 The Structure of BglII Bound to its Cognate DNASite 227Conclusions 232References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 233

XIV

Structure and Function of the Tetrameric Restriction Enzymes

V. SIKSNYS, S.GRAZULIS, R . HUBER

Contents

237

1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . 2372 Structural Anatomy of the Tetrameric Restriction Enzymes 2392.1 Monomer Structure . . 2392.2 Dimer Arrangement 2412.3 Tetramer Organization 2433 Active Sites of the Tetrameric Restriction Enzymes 2464 DNA Recognition by NgoMIV Restriction Endonuclease 2505 Possible Model of DNA Recognition

by Bse634I/Cfr10I Restriction Endonucleases . .. . . 2536 Functional Significance of the Tetrameric Architecture

of Restriction Enzymes 256References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 258

Structure and Function of Type lIE Restriction Endonucleases - or:From a Plasmid that Restricts Phage Replication to a NewMolecular DNARecognition Mechanism . . . . . . . . . . . . . . . . 261

M. REUTER, M. MUCKE, D.H. KRUGER

12

3

4

5

5.15.25.3

678

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . .The General Problem: Type lIE REases Needthe Simultaneous Interaction with Two Copiesof Their Recognition Sequence for Enzymatic ActivityActivation of Type lIE REases by SyntheticOligonucleotide Duplexes .Activation of Type lIE REases by Cleavage Productsand by Non-Cleavable Oligonucleotide DuplexesThe Enzymes' Reaction Mechanism -General Aspects and Details .Cooperative Interaction with Two Recognition Sites . . . .Stoichiometry of the Active Enzyme-Substrate ComplexesHow Do Type lIE REases Communicate BetweenRemote DNA Recognition Sites in a DNA Molecule? . . .Sequence-Specific DNA Recognition by Type lIE REasesDomain Organization of Type lIE REases . . . . . . . . .Modular Architecture of NaeI and EcoRIIand Its Functional Implications .

261

262

265

265

267267268

270272

275

278

Contents XV

9 Reaction Mechanism of Type lIE REases ..... 28010 Are NaeI and EcoRII Evolutionary Links Between

REases,Topoisomerases and DNARecombinases? 28311 Does Nature Construct Proteins with New Functions

by Shuffling Protein Domains? . . . . . . . . . . . . . . 28512 Activation of Type lIE REases for Biotechnological

Purposes and for Mapping Epigenetic DNAModifications . 28713 Final Remark 288References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 289

Analysis of Type II Restriction EndonucleasesThat Interact with TwoRecognition Sites

A.T. WELSH, S.E. HALFORD, D.T. SCOTT

1 Introduction . . . . . . . .2 Different Classes of Interactions3 Information from cisReactions3.1 One-Site/Two-Site Assays .3.2 Determining Reaction Mechanism3.3 The Effect of Supercoiling ... ..3.4 Catenane Substrates . . . . . . . . .4 DNABinding and Looping with Sites in cis5 Information from trans Reactions5.1 Kinetic Studies5.2 Binding Studies6 ConclusionReferences . . . . . . . . .

The Role of Water in the EcoRI-DNA Binding

N. SIDOROVA, D.C. RAU

297

297298301301302306306307311311312314315

319

1233.13.1.1

Introduction . . . . . . . . . .Thermodynamics . . . . . . .Experimental Applications . .Equilibrium Competition . . .Osmotic Stress Dependence or Knonsp-sp

319321325325325

XVI Contents

3.1.2 pH and Salt Dependence of Knonsp-sp . . . . . . . . . 3263.2 Dissociation Kinetics of EcoRI from Its Specific Site 3273.2.1 Osmotic Dependence of k, . . . . . . . . . . . . . . 3283.2.2 pH and Salt Dependence of kd • • • • • • • • • • • • 3293.3 Removing Water from an EcoRI-Noncognate DNA

Complex with Osmotic Stress 3303.3.1 Competitive Equilibrium at High Osmotic Stress 3313.3.2 Dissociation Kinetics of the EcoRI from Noncognate Sites 3323.4 Other Applications of Osmotic Stress

to Restriction Nucleases 3333.5 Application of Hydrostatic Pressure

to Restriction Nucleases 3334 Summary . 334References . . . . . . . . . . . . . . 335

Role of Metal Ions in Promoting DNABindingand Cleavage by Restriction Endonucleases

I.A.COWAN

1 Introduction . . . . . . . . . . . . . . . .2 Selection of Metal Cofactors to Promote

Endonuclease Activity .3 Magnesium Analogs .4 Inhibitory Influence of Metal Cofactors .5 Illustrative Examples and General Guidelines

for Metal-Promoted Endonuclease Activity5.1 EcoRV and EcoRI5.2 Pvull .5.3 BamHI .6 Other Restriction Endonucleases7 Concluding RemarksReferences . . . . . . . . . . . . . . . . . . .

339

339

341342342

343347350353355356357

Contents

Restriction Endonudeases: Structure of the ConservedCatalytic Core and the Role of Metal Ions in DNACleavage

J.R. HORTON, R.M. BLUMENTHAL, X.CHENG

XVII

361

1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . 3612 Common Structural Attributes of Type II REases 3653 A Common Catalytic Core with Key Catalytic Sidechains 3654 Phylogenetic Analysis of REases . . . 3675 Possible Roles for Divalent Metal Ions . . . 3686 The One-Two-Three Metal Debate . . . . . 3697 Generalizations Regarding REase Catalysis . 3708 One-Metal Catalytic Mechanism: EcoRI, BglII 3759 Two-Metal Catalytic Mechanisms 3779.1 EcoRV. . . . . . 3789.2 BamHI . . . . . . 3799.3 Tn7 Transposase . 3809.4 T7 Endonuclease I 38010 Three-Metal Catalytic Mechanism 38111 "Hypothetical" Active Sites of REase Superfamily Members 38211.1 MunI 38211.2 BsoBI 38311.3 NaeI 38311.4 MutH 38411.5 Holiday Junction Resolvases, Hjc 38512 Prospects for the Future 385References . . . . . . . . . . . . . . . . . . . 386

Protein Engineering of Restriction Enzymes

J.ALVES, P.VENNEKOHL

393

1 Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . 3932 Modifying Contacts of Restriction Enzymes

to Their Recognition Sequence 3932.1 Site-Directed Mutagenesis 3972.2 In Vivo Selection Systems . . . . . . . . 4003 Lengthening of Recognition Sequences 4024 Changing the Subunit Composition . . 4055 Future Directions (In Vitro Evolution) 406References . . . . . . . . . . . . . . . . . . . . . . 407

XVIII

Engineering and Applications of Chimeric Nucleases

K. KANDAVELOU, M . MANl, S. DURAl, S. CHANDRASEGARAN

Contents

413

1 Introduction . . . . . . . . . . . . . . . . . . . . . 4132 Engineering of Chimeric Nucleases . . . . . . . . 4142.1 Functional Domains in FokI Restriction Endonuclease 4162.2 Chimeric Nucleases . . . . . . . . . . . . . . . . . . 4162.3 Zinc Finger Binding and Specificity . . . . . . . . 4172.4 Mechanism of Cleavageby Zinc Finger Chimeric

Nucleases (ZFN) . . . . . . . . . . . . . . . . 4223 Application of Chimeric Nucleases 4233.1 Stimulation of Homologous Recombination

Through Targeted Cleavage in Frog Oocytes Using ZFN:Recombinogenic Repair 423

3.2 Targeted Chromosomal Cleavage and Mutagenesisin Fruit Flies Using ZFN:Mutagenic Repair. 424

3.3 Gene Targeting in Human Cells Using ZFN . . . . . 4254 Future Experiments . . . . . . . . . . . . . . . . . . 4264.1 Targeted Chromosomal Cleavage and Mutagenesis

of the CCR5 Gene in a Human Cell Line 4274.2 Targeted Correction of a CFTR Genetic Defect

in a Human Cell Line Using ZFN . . . . . . . . 4285 Potential Limitations of the Chimeric Nuclease Technology 4296 Future Outlook 430References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 431

Subject Index 435

Contributors

AGGARWAL, ANEEL K.

Structural Biology Program, Department of Physiology and Biophysics,Mount Sinai School of Medicine, 1425 Madison Ave,New York, New York10029, USA

ALVES, JURGEN

Department of Biophysical Chemistry, Medical School Hanover,Carl -Neuberg-Str, 1,30625 Hanover, Germany

BELFORT, MARLENE

Molecular Genetics Program, New York State Department of Health,Albany, New York 12201-2002, USA

BESTOR, TIMOTHY

Genetics and Development, Columbia University, New York, New York10032, USA

BHAGWAT, ASHOK S.

Department of Chemistry,Wayne State University, Detroit, Michigan48202, USA

BICKLE, THOMAS A .

Department of Microbiology, Biozentrum, Universitat Basel, 4056 Basel,Switzerland

BITINAITE, JURATE

New England Biolabs , 32 Tozer Road, Beverly, Massachusetts 01915, USA

xx Contributors

BLUMENTHAL, ROBERT M.

Department of Microbiology & Immunology, and Program inBioinformatics & ProteomicslGenomics, Medical College of Ohio, Toledo,Ohio 43614-5806, USA

BUJNICKI, JANUSZ M.

Bioinformatics Laboratory, International Institute of Molecular and CellBiology in Warsaw, Trojdena 4, 02-109 Warsaw, Poland

CHANDRASEGARAN, SRINIVASAN

Department of Environmental Health Sciences, The Johns HopkinsUniversity Bloomberg School of Public Health, 615 North Wolfe Street,Baltimore, Maryland 21205-2179, USA

CHANDRASEKHAR, K.

Department of Biological Sciences, University of Pittsburgh; Pittsburgh,Pennsylvania 15260,USA

CHENG, XIAODONG

Department of Biochemistry, Emory University School of Medicine, 1510Clifton, Atlanta, Georgia 30322, USA

COWAN,J.A.

Evans Laboratory of Chemistry, The Ohio State University, 100 West 18thAvenue, Columbus, Ohio 43210, USA

DEGTYAREV, SERGEY K.

SibEnzyme, 630090 Novosibirsk, Russia

DRYDEN, DAVID T.F.

School of Chemistry, University of Edinburgh, The King's Buildings,Edinburgh EH9 3JJ,UK

DURAl, SUNDAR


DYBVIG, KEVIN

Department of Genetics, University of Alabama at Birmingham,Birmingham, Alabama 35294, USA

Contributors

FIRMAN, KEITH

Biophysics Laboratories, School of Biological Sciences, University ofPortsmouth, Portsmouth PO1 2DT, UK

GRAZULIS, SAULIUS

Institute of Biotechnology, Graiciuno 8,Vilnius 2028, Lithuania

XXI

GRIGORESCU, ARABELA


GROMOVA, ELIZAVETA S.

A.N. Belozersky Institute of Physico-Chemical Biology, Moscow StateUniversity, 119992 Moscow, Russia

GUMPORT, RICHARD 1.

The University of Illinois College of Medicine, Urbana, Illinois61801-3602, USA

HALFORD, STEPHEN E.

Department of Biochemistry, School of Medical Sciences, University Walk,University of Bristol, Bristol BS8 1TD, UK

HATTMAN, STANLEY

Department of Biology, University of Rochester, Rochester, New York14627-0211, USA

HEITMAN, JOSEPH

Howard Hughes Medical Institute, Duke University Medical Center,Durham, North Carolina 27710, USA

HORNBY, DAVID P.

Department of Molecular Biology and Biotechnology, University ofSheffield, Firth Court, Western Bank, Sheffield S10 2TN, UK

HORTON, JOHN R .

Department of Biochemistry, Emory University School of Medicine, 1510Clifton, Atlanta, Georgia 30322, USA

XXII Contributors

HORVATH, MONICA


HUBER, ROBERT

Max Planck Institute of Biochemistry, Am Klopferspitz 18a, 8215Martinsried, Germany

JANULAITIS, ARVYDAS

Institute of Biotechnology, 2028 Vilnius, Lithuania

JELTSCH, ALBERT

Institut fur Biochemie, FB 8, Heinrich-Buff-Ring 58, [ustus-LiebigUniversitat, 35392 Giessen, Germany

JOSEPHSEN, JYTTE

Department of Dairy and Food Science, Royal Veterinary and AgriculturalUniversity, 1958 Frederiksberg C, Denmark

KANDAVELOU, KARTHIKEYAN


KISS,ANTAL

Institute of Biochemistry, BRC,6701 Szeged, HungaryKlaenhammer

KOBAYASHI, ICHIZO

Institute of Medical Science, University of Tokyo, 4-6-1, Shirokanedai,Minato-ku, Tokyo 108-8639, Japan

KONG, HUIMIN

New England Biolabs, 32 Tozer Road, Beverly,Massachusetts 01915, USA

KRUGER, DETLEV H .

Institut fur Virologie, Universitatsklinikum Charite,Humboldt-Universitat, Schumannstr. 20121,10098 Berlin , Germany

LACKS, SANFORD

Brookhaven National Laboratory, Upton, New York 11973-5000, USA

Contributors XXIII

LUKACS, CHRISTINE M.

Structural Biology Program, Department of Physiology and Biophysics,Mount Sinai School of Medicine, 1425 Madison Ave, New York,New York 10029, USA

MANI,MALA


MARINUS, MARTIN G.

Department of Pharmacology, University of Massachusetts MedicalSchool, Worcester, Massachusetts 01655, USA

MCCLELLAND, SARAH E.

DNA-Protein Interactions Group, Department of Biochemistry, School ofMedical Sciences, University of Bristol, Bristol, BS8 1TD, UK

MIYAHARA, MICHIKO

National Institute of Health Sciences, 1-18-1, Kamiyoga, Setagaya-ku,Tokyo 158-8501, Japan

MORGAN, RICHARD D.

New England Biolabs, 32 Tozer Road, Beverly, Massachusetts 01915, USA

MUCKE, MERLIND

Institut fur Virologie, Universitatsklinikum Charite,Humboldt-Universitat, Schumannstr. 20/21,10098 Berlin, Germany

MURRAY, NOREEN E.

Institute of Cell and Molecular Biology, University of Edinburgh, TheKing's Buildings, Edinburgh EH9 3JR, UK

NAGARAJA, VALAKUNJA

Department of Microbiology and Cell Biology, Indian Institute of Science,560012 Bangalore, India

PIEKAROWICZ, ANDRZEJ

Institute of Microbiology, Warsaw University, Miecznikowa 1,02-096Warsaw, Poland

XXIV Contributors

PINGOUD, ALFRED

Institut fur Biochemie, Justus-Liebig- Universitat, 35392 Giessen, Germany

PROTA, ANDREA E .

Biomolecular Research, Paul Scherrer Institut, 5232 Villigen, Switzerland

RALEIGH, ELISABETH


RAO, DESIRAZU N.

Department of Microbiology and Cell Biology, Indian Institute of Science,560012 Bangalore, India

RAU, DONALD C.

Laboratory of Physical and Structural Biology, National Institute of ChildHealth and Human Development, National Institutes of Health, BId. 9,Room 1E108, Bethesda, Maryland 20892, USA

REICH, NORBERT

University of California, Santa Barbara, Santa Barbara, California93106-0001, USA

REPIN, VLADIMIR E .

State Research Center of Virology and Biotechnology'Vektor', Koltsovo,Novosibirsk Region 633059, Russia

REUTER, MONIKA

Institut fur Virologie, Universitatsklinikum Charite,Humboldt-Universitat, Schumannstr. 20/21,10098 Berlin, Germany

ROBERTS, RICHARD J.


ROSENBERG, JOHN M.

Department of Biological Sciences, University of Pittsburgh; Pittsburgh,Pennsylvania 15260, USA

SCHEURING, VANAMEE EVA

Structural Biology Program, Department of Physiology and Biophysics,Mount Sinai School of Medicine, 1425 Madison Ave, New York, New York10029, USA

Contributors xxv

SCOTT, DAVID J.Department of Biochemistry, School of Medical Sciences, University Walk,University of Bristol, Bristol BS8 ITD, UK

SELKER, ERIC U.

Institute of Molecular Biology, University of Oregon, Eugene, Oregon97403, USA

SHAW, PANG-CHUI

Department of Biochemistry, The Chinese University of Hong Kong, HongKong

SIDOROVA, NINA

Laboratory of Physical and Structural Biology, National Institute of ChildHealth and Human Development, National Institutes of Health, BId. 9,Room IEI08, Bethesda, Maryland 20892, USA

SIKSNYS, VIRGINIJUS

Institute of Biotechnology, Graiciuno 8,Vilnius 2028, Lithuania

STEIN DANIEL D .

Department of Microbiology, University of Maryland, College Park,Maryland 20742, USA

STODDARD, BARRY L.

Fred Hutchinson Cancer Research Center, Seattle, Washington 98109, USA

SZCZELKUN, MARK D .

DNA-Protein Interactions Group, Department of Biochemistry, School ofMedical Sciences, University of Bristol, Bristol, BS8 ITD, UK

SZYBALSKI, WACLAW

McArdle Laboratory, University of Wisconsin, Madison, Wisconsin 53706,USA

TODD R.

Departments of Food Science and Microbiology, North Carolina StateUniversity, Raleigh, North Carolina 27695-7624, USA

XXVI Contributors

TRAUTNER, THOMAS A.

MPI fur Molekulare Genetik, Ihnestrasse 73, 14195 Berlin, Germany

URBANKE, CLAUS

Zentrale Einrichtung Biophysikalisch-Biochemische Verfahren,Medizinische Hochschule, 30625 Hannover, Germany

VAN ETTEN, JAMES L.

Department of Plant Pathology, University of Nebraska-Lincoln, Lincoln,Nebraska 68583, USA

VENNEKOHL, PETRA

Department of Biophysical Chemistry, Medical School Hanover,Carl-Neuberg-Str, 1,30625 Hanover, Germany

VIADIU, HECTOR

Structural Biology Program, Department of Physiology and Biophysics,Mount Sinai School of Medicine, 1425 Madison Ave,New York, New York10029, USA

VITOR, JORGE M.B.

Faculdade de Farmacia de Lisboa, 1600-083 Lisbon, Portugal

WELSH, ABIGAIL J.

Department of Biochemistry, School of Medical Sciences, University Walk,University of Bristol, Bristol BS8 1TD, UK

WILKOSZ, PATRICIA A.

Department of Biological Sciences, University of Pittsburgh; Pittsburgh,Pennsylvania 15260, USA

WILSON, GEOFFREY G.


WINKLER, FRITZ K.

Biomolecular Research, Paul Scherrer Institut, 5232 Villigen, Switzerland

XU,SHUANG-YONG


nucleic acids andmolecular biology 14978-3-642-18851-0/1.pdfnucleic acid interactions, e.g., how...

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