2013 doi ?-amylase an enzyme specificity found in various families of glycoside hydrolases

7/29/2019 2013 DOI ?-Amylase an Enzyme Specificity Found in Various Families of Glycoside Hydrolases

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DOI 10.1007/s00018-013-1388-z Cellular and Molecular Life SciencesCll. Mol. Li Sci.

RevIew

Amylase: an enzyme specifcity ound in various amilieso glycoside hydrolases

tean Janecek Birte Svensson E. Ann MacGregor

Rcid: 26 April 2013 / Risd: 27 May 2013 / Accptd: 27 May 2013 Springr Basl 2013

amily GH57 attributs ar likly to b charactristic o-amylass rom th amily GH119, too. with rgard toamily GH126, conrmation o th unambiguous prsnco th -amylas spcicity may nd mor biochmi-cal instigation bcaus o an obious, but unxpctd,homology ith inrting -glucan-acti hydrolass.

Keywords -Amylas Glycosid hydrolas amilis GH13, GH57, GH119, GH126 Consrd squncrgions Catalytic machinry eolutionary rlationships

AbbreviationsCAZy Carbohydrat-Acti nZyms

CBM Carbohydrat-binding modulCSR Consrd squnc rgionGH Glycosid hydrolasSBD Starch-binding domainTIM Trios-phosphat isomras

Introduction

-Amylas rprsnts th bst knon and most intn-sily studid amylolytic nzym. Amylolytic nzyms arnzyms dgrading starch and starchy substrats and arapplid idly in arious branchs o th ood, pharmacu-tical, and chmical industris. In a broadr sns, th dsig-nation amylolytic nzyms has bn usd or th arityo starch hydrolass and rlatd nzyms that ar actiin trms o hydrolysis, transglycosylation, and isomriza-tiontoards th -glucosidic bonds prsnt in starch andrlatd poly- and oligo-saccharids.

As our undrstanding o th dtails o protin primaryand trtiary structur o indiidual amylolytic nzymsincrasd in th last coupl o dcads, it has bcom

Abstract -Amylas (eC 3.2.1.1) rprsnts th bst

knon amylolytic nzym. It catalyzs th hydrolysis o-1,4-glucosidic bonds in starch and rlatd -glucans. Ingnral, th -amylas is an nzym ith a broad substratprrnc and product spcicity. In th squnc-basdclassication systm o all carbohydrat-acti nzyms,it is on o th most rquntly occurring glycosid hydro-lass (GH). -Amylas is th main rprsntati o amilyGH13, but it is probably also prsnt in th amilis GH57and GH119, and possibly n in GH126. Family GH13,knon gnrally as th main -amylas amily, orms clanGH-H togthr ith amilis GH70 and GH77 that, ho-r, contain no -amylas. within th amily GH13, th-amylas spcicity is currntly prsnt in sral sub-amilis, such as GH13_1, 5, 6, 7, 15, 24, 27, 28, 36, 37,and, possibly in a mor that ar not yt dnd. Th-amylass classid in amily GH13 mploy a ractionmchanism giing rtntion o conguration, shar 47consrd squnc rgions (CSRs) and catalytic machin-ry, and adopt th (/)8-barrl catalytic domain. Althoughth amily GH57 -amylass also mploy th rtainingraction mchanism, thy possss thir on CSRs andcatalytic machinry, and adopt a (/)7-barrl old. Ths

. Janck (*)Laboratory o Protin eolution, Institut o MolcularBiology, Sloak Acadmy o Scincs, Dbrask csta 21,84551 Bratislaa, Sloakia-mail: [email protected]

B. Snssonenzym and Protin Chmistry, Dpartmnt o Systms Biology,Th Tchnical Unirsity o Dnmark, Sltots Plads, Building224, 2800 Kgs. Lyngby, Dnmark

e. A. MacGrgor2 Nicklaus Grn, Liingston, wst Lothian eH54 8RX, UK


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clar that ths nzyms, hich ar ry closly rlatd bythir unction on starch, .g., -amylas, -amylas, andglucoamylas, ar not closly rlatd in trms o struc-tur and raction mchanism. It has turnd out to b morappropriat to bas a classication o amylolytic nzymson similaritis in thir amino acid squncs and thr-dimnsional structurs, raction mchanisms, and catalytic

machinris, all rfcting olutionary rlatdnss, than onspcicity. Such an approach, hor, opns th door toidas that nzyms closly rlatd in unction ill classiysparatly, but also that similar ractions can b catalyzdby structurally dirnt and thus olutionarily unrlatdprotins.

This appars to b th situation or th -amylasnzyms that ar th topic o th prsnt ri. Thmain goal is to dscrib th most rcnt knoldg o-amylass ith rgard to th xistnc o structurally di-rnt nzyms apparntly possssing th sam -amylas-typ actiity. Sinc th -amylas is an nzym ith a

broad substrat prrnc and product spcicity, mpha-sis is also gin to subtl but uniqu structural aturs dis-criminating btn closly rlatd -amylass, taking intoaccount, or xampl, thir taxonomical origin.

CAZy classifcation system and amylases

-Amylass ar glycosid hydrolass (GHs) and hathror bcom a part o th squnc-basd classica-tion systm o nzyms acti toards arious saccharids.This systm as rst dlopd som 20 yars ago or GHs

[1] and latr updatd [2, 3]. No, th ntir systm xistsonlin (sinc 1998) or all Carbohydrat-Acti nZyms(CAZy), as th so-calld CAZy databas (or CAZy srr;http://.cazy.org/) coring: (1) catalytic modulsinold in dgradation, cration, and modication o gly-cosidic linkags o saccharids; and (2) associatd carbo-hydrat-binding moduls (CBM) rsponsibl or adhsionto saccharids [4]. Th classication o catalytic modulsincluds, in addition to GHs, glycosyl transrass, poly-saccharid lyass, and carbohydrat strass.

Th CAZy classication systm basd on comparisono amino acid squncs as stablishd in an ort toorcom th inability o classical IUB enzym Nomn-clatur (or GHs: eC 3.2.1.x) to rfct structural atursand olutionary aspcts o GHs [1]. within th CAZysystm, indiidual nzyms ar classid into squnc-basd amilis. Currntly, mor than 130 such amilis(dsignatd ith Arabic numrals, .g., GH13) ha bncratd chronologically sinc 1991. Th nzyms (pro-tins) blonging to th sam GH amily should, in princi-pl, xhibit squnc similaritis (usually ith consrdsqunc rgions; CSRs), shar catalytic machinry (th

sam catalytic rsidus locatd on corrsponding struc-tural lmnts), mploy th sam raction mchanism(ithr rtaining or inrting) and adopt th sam typ ocatalytic domain old [1]. Thr ar to additional lls ohirarchical classication in th CAZy databas, i.., clansas a highr ll and subamilis as a lor ll o hir-archy [4, 5]. A GH clan groups togthr amilis sharing

catalytic machinry and adopting th sam structural oldo th catalytic domain, but ith signicant dirnc inorall squnc. A clan is dsignatd by a lttr prcddby a dash (.g., GH-H) attributd in th alphabtical ordro thir apparanc, ith ry GH clan cration bingbasd on trtiary structur inormation [4]. A GH subam-ily is a group o mmbrs o on amily that shars morsqunc/structural and unctional charactristics thanapplicabl or th ntir amily, i.., a subamily mmbrsshould shar a mor rcnt olutionary ancstor. Subami-lis o a gin GH amily ar dsignatd by a numral su-x prcdd by undrscor (.g., GH13_7). In th scintic

litratur proposals ha bn mad to split arious GHamilis into subamilis, and th -amylas amily GH13as th rst to ha bn ocially diidd in this ay, inthis cas into 37 subamilis, by th CAZy curators [6]. Inaddition to GH13, o all 131 GH amilis, ocial subami-lis ha bn dnd only or th amilis GH5 [7] andGH30 [8]. Moror, th ntris or th amilis GH21,GH40, GH41, GH60, and GH69 r dltd rom th GHsystm (CAZy; [4]).

Currntly, th ntir CAZy databas cors th squncdata rom mor than 2,100 gnoms o Bactria, almost150 gnoms o Archaa and around 70 o eukaryots

(CAZy; [4]). Th ncyclopdic projct CAZypdia (http://.cazypdia.org/) as cratd about 5 yars ago and,bing rittn and curatd by scintists dirctly inold instudying such nzyms, it constituts a ry comprhnsiand complmntary sourc o inormation.

-Amylas (eC 3.2.1.1), th most idly studid amylo-lytic nzym, catalyzs th hydrolysis, ith rtntion oconguration, o th intrnal -1,4-glucosidic bonds instarch and rlatd poly- and oligo-saccharids and is anndo-acting nzym [9]. Som xo-acting nzyms can bconsidrd as -amylass, but ar not addrssd in dtailhr. Th -amylass, hor, xhibit quit arid prod-uct prols, dpnding on thir origin [10]. In addition toth main hydrolytic raction, many -amylass ar alsoabl to catalyz transglycosylation, .g., thos rom Asper-gillus oryzae [11], Alteromonas haloplanktis [12], humansalia [13], and human pancras [14], and n a chimraromBacillus amyloliqueaciens andBacillus licheniormis[15]. without a srious biochmical charactrization, th-amylas spcicity should not b ascribd unambigu-ously to an nzym displaying simply amylolytic (i.., astarch-hydrolyzing) actiity [16]. Any xprimntal data,
http://www.cazy.org/http://www.cazypedia.org/http://www.cazypedia.org/http://www.cazypedia.org/http://www.cazypedia.org/http://www.cazy.org/


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-Amylase within the CAZy classifcation system

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on th othr hand, can no b supportd by rliabl in sil-ico analyss o primary structur and prdictions.

Currntly, th -amylas nzym spcicity ithin thCAZy databas can b ound unambiguously in th ami-lis GH13, GH57 and GH119, moror it has bn sug-gstd to also b prsnt in amily GH126 (CAZy; [4, 17].O ths, th amily GH13 can b considrd to b th

main -amylas amily rst proposd in th CAZy systmin 1991 [1], ith amily GH57 as th scond and smallr-amylas amily stablishd in 1996 [3] and th amilisGH119 and GH126 dnd in 2006 and 2011, rspctily(CAZy; [4]).

Amylase amily GH13

-Amylas amily GH13 as stablishd in 1991 hnth classication o GHs, i.., th CAZy databas [4], asrst publishd [1]. At that tim, th amily GH13 as (du

to classication critria) dnd as a polyspcic nzymamily coring, in addition to -amylas, nzyms suchas -glucosidas, dxtran glucosidas, isoamylas, pul-lulanas, amylopullulanas, nopullulanas, cyclodxtringlucanotransras, and som xo-acting amylass [1].Th classication rfctd th prdictions mad pri-ously that arious starch hydrolass and rlatd amylolyticnzyms ould xhibit mutual squnc similaritis andshar catalytic rsidus and olds [1821]. Currntly, thamily GH13 contains approximatly 30 dirnt nzymspcicitis rom thr eC groups, i.., hydrolass, trans-rass, and isomrass [4, 22], plus non-nzymatic mm-

brs rprsntd by th hay-chains o htromric aminoacid transportrs and 4F2 antigns [2325]. with rgard toth numbr o squncs blonging to GH13, this amilyranks among th largst in CAZy ith mor than 13,500mmbrs in March 2013 (CAZy; [4]) originating prdomi-nantly rom Bactria (~11,500), ith r rom eukary-ots (~1,800) and Archaa (~200).

In gnral, th GH13 -amylass and othr amily GH13mmbrs ar thr-domain protins (Fig. 1) consisting oth main catalytic (/)8-barrl domain (domain A) itha small domain B protruding out o th barrl as a longrloop btn th strand 3 and hlix 3 and succdd atth C-trminal nd by domain C, adopting an antiparal-ll -sandich old [10, 2630]. This domain organiza-tion as rst dtrmind or Taka-amylas A [31], i.., th-amylas romA. oryzae. Th domain o th (/)8-barrlis composd o ight innr paralll -strands surrounddby ight -hlics and, bcaus it as rst rcognizd inth structur o trios-phosphat isomras (TIM; [32]), itis otn calld th TIM-barrl [33]. Indiidual GH13 mm-brs and somtims all mmbrs ith a gin spcic-ity may contain additional domains on ithr trminus o

thir polypptid chain. Although thir unctions ha stillnot bn compltly undrstood, such domains ar otninold in binding starch, glycogn, and othr rlatd sac-charids [3438]. Typical starch-binding domains (SBDs)ha also bn classid ithin th CAZy databas as thso-calld CBM amilis [4] ith CBM20 as a rprsnta-ti o th C-trminal SBD that as rst rcognizd [39

41]. At prsnt, tn CBM amilis ar considrd as SBDamilis: CBM20, CBM21, CBM25, CBM26, CBM34,CBM41, CBM45, CBM48, CBM53, and CBM58 [4245].

History

All th amily GH13 mmbrs should oby sral distinctcritria [10, 46]: (1) mploying th rtaining mchanism o-glycosidic bond claag; (2) adopting a (/)8-barrl asth catalytic domain; (3) xhibiting 47 CSRs positiondmostly on -strands o th barrl (Fig. 2); and (4) sharingth catalytic machinry, consisting o strand 4-aspartic

acid (catalytic nuclophil), 5-glutamic acid (protondonor), and 7-aspartic acid (transition-stat stabilizr)hr rrrd to as th catalytic triad. Ths critria rdnd xplicitly by [47] in a mor stringnt rsion thatrfctd th situation 20 yars ago hn a much lornumbr o squncs as aailabl.

Throughout amily GH13, squnc idntity isxtrmly lo and only th catalytic triad, i.., Asp206,Glu230, and Asp297 (Taka-amylas A numbring; [31]),plus th arginin (Arg204) positiond to rsidus borth catalytic nuclophil (Fig. 2) ar consrd inariantly[46]. This is, hor, not applicabl or th non-nzymatic

GH13 mmbrs, that, dpnding on thir taxonomic origin,may not contain th catalytic rsidus [23, 25].

In gnral, hor, most unctionally important andothr consrd rsidus or any GH13 amily mmbr around in th 47 CSRs [46] (Fig. 2). Th our bst knonrgions, i.., CSRs I, II, III, and Iv, r ll stablishdin 1986 by comparison o 11 -amylass originating rommicroorganisms, plants and animals [48]. It is orth mn-tioning that th CSRs I, II, and Iv r initially proposdby Toda t al. [49] ho pointd out ths rgions or Taka-amylas A and pig pancratic -amylas, thn by Fridbrg[50] ho addd th rgions in th B. amyloliqueaciens-amylas and nally by Rogrs [51] ho compltd thpictur by dscribing thm in th barly -amylas. Ththr additional rgions, i.., CSRs v, vI, and vII, ridntid latr [52, 53]. Th rst our CSRs ar locatd ator nar th C-trmini o strands 3, 4, 5 and 7 o thcatalytic (/)8-barrl domain and includ th catalytictriad. Th thr additional CSRs, positiond nar th C-tr-minus o domain B and at or nar th C-trmini o th bar-rl strands 2 and 8, contain rsidus that may b usd todistinguish th GH13 spcicitis rom ach othr.


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Clan GH-H

Noadays, nzyms rlatd structurally to -amylas arrprsntd by th CAZy clan GH-H consisting o thramilis, i.., th amilis GH70 and GH77 in addition toth main amily GH13 [4, 10].

Th amily GH70 contains glucosyltransrass hainga circularly prmutd rsion o th -amylas-typ (/)8-barrl catalytic domain, rst prdictd by MacGrgor t al.[54] and conrmd by soling th thr-dimnsional struc-tur oLactobacillus reuteri glucansucras [55] ollodby th structurs o a closly rlatd nzyms [5658].

Th mmbrs o amily GH77 ar 4--glucanotransrassith a rgular catalytic (/)8-barrl, but lacking thdomain C that ollos th barrl in amily GH13 [5961].Moror, in sral GH77 4--glucanotransrass rom

Borrelia, th abo-mntiond consrd arginin that issituatd to rsidus prcding th catalytic nuclophil,

in CSR II, is substitutd by a lysin [62]. This mans that,hn th nzyms o clan GH-H ar considrd as a hol,only th catalytic triad is truly inariantly consrd [63].

Dspit th obsrd dirncs btn th indiidualGH amilis o clan GH-H, thr is no doubt that all thmmbrs o this clan containing -amylass (i.., GH13,GH70, and GH77) shar a common ancstor [64, 65] andmay b radily discriminatd rom th rmotly homolo-gous amily GH31 o-glucosidass [66].

Subamilis in GH13

Th many spcicitis, larg numbr o squncs, andobious subgroups o nzyms, .g., th so-calld oligo-1,6-glucosidas and nopullulanas subamilis [67], sug-gstd th nd or urthr subdiision o GH13. A majorbrakthrough to dscrib th amily mmbrs at a lorll o hirarchy cam in 2006 hn GH13 as brokn

(a) (b) (c)

(d) (e) (f)

(g)

Fig. 1 Trtiary structurs o rprsntati amily GH13 -amylass.Th structurs rom olloing subamilis and origins ar shon: aGH13_1, Aspergillus oryzae (PDB cod: 2TAA; [31]); b GH13_5,Bacillus licheniormis (PDB cod: 1BLI; [249]); c GH13_6, Hor-deum vulgarebarly isozym AMY-1 (PDB cod: 1P6w; [129]); dGH13_37, unculturd bactrium AmyP (modld structur; rsidusLu6-Thr491) obtaind rom th Phyr srr [245] basd on th Fla-vobacterium sp. 92 GH13 cyclomaltodxtrinas (PDB cod: 3eDe;[205]) as tmplat; e unclassid,A. haloplanktis (PDB cod: 1G94;[12]); unclassid, Halothermothrix orenii AmyB (PDB cod:

3BC9; [70]); g unclassid, Bacteroides thetaiotaomicron (PDBcod: 3K8L; [38]). Th indiidual domains ar colord as ollos:catalytic (/)8-barrl blue, domain B green, domain C red, N-trmi-

nal domain cyan; starch-binding domain o CBM58 amily magenta.Th GH13 catalytic triad, i.., catalytic nuclophilAsp ( top), pro-ton donorGlu (let) and transition-stat stabilizrAsp (right)ishighlightd in th (/)8-barrl domain in ach structur in a similarposition. Saccharid molculs ar colored yellow to mphasiz: c anadditional surac binding sit as a pair o sugar tongs ith th sid-chain o tyrosin (in black) inold in binding; e a hptasaccharidoccupying th subsits rom 4 to +3 as a transglycosylation prod-uct rom acarbos (a psudottrasaccharid); and g a maltopntaosbound by th SBD o th amily CBM58 insrtd ithin th domain

B. Th structurs r rtrid rom th Protin Data Bank (PDB;[250]) and isualizd ith th program wbLabvirLit (Molcu-lar Simulations, Inc.)


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up into 35 subamilis by CAZy curators [6]. This basicallyrfcts th ida that thr ar groups o nzyms in amilyGH13 that xhibit, ithin such a subamily, a substantiallyhighr dgr o similarity in squnc, taxonomy, and/orspcicity than in amily GH13 as a hol. Currntly, 37subamilis o GH13 ha bn dnd (CAZy; [4]), butsral squncs and charactrizd nzyms ar not yt

assignd to a subamily [38, 6871].

Subamily GH13_1

Th subamily GH13_1 cors th ukaryotic -amylassrom ungi and yast only [6] ith no -amylas o a di-rnt taxonomic origin (CAZy; [4]). Th numbr 1or this GH13 subamily may ll rfct th act that th-amylas rom A. oryzae (i.., Taka-amylas A; [49]) isclassid hr, hich as th rst -amylas ith its thr-dimnsional structur sold (Fig. 1; [31]). Trtiary struc-turs ar aailabl also or to additional closly rlatd-amylass romAspergillus niger; on bing th so-calldacid-stabl -amylas [72] and th othr on xhibiting100 % squnc idntity to Taka-amylas A [73]. Thr arto additional Taka-amylas A trtiary structurs xhibit-ing an incrasd thrmostability, on ith chmically mod-id Asp197 [74] and th othr as a hay-atom driati[75].

Although thr is no structur as yt or GH13_1-amylas rom a yast (CAZy; [4]), sral yast-amylass r dmonstratd to possss SBDs o ami-lis CBM20, .g., th nzyms rom Cryptococcus sp. S-2[76] and Cryptococcus avus [77], or CBM21, .g., th

nzyms romLipomyces kononenkoae [78] andLipomycesstarkeyi [79]. Som sho an ability to dgrad ra starchithout a distinct SBD, or xampl th -amylas oSac-charomycopsis fbuligera KZ [80]. This may rfct thprsnc o surac (i.., scondary) binding sits, hich isa gnral atur o CAZy [81]. Approximatly hal o thknon surac binding sits ha bn ound ithin sralGH13 subamilis [82], but thy ar bst knon rom bar-ly and othr plant -amylass o subamily GH13_6 [83].In th subamily GH13_1, a surac binding sit as sn,.g., outsid th acti sit in th A. niger-amylas struc-tur [73]. whil SBDs may b prsnt also in som un-gal -amylass, .g., rom Aspergillus kawachii [84], it isnot clar hy som o ths -amylass contain an SBDhras othrs do not [40, 41, 85].

Intrstingly, th GH13_1 mmbrs xhibit simi-laritis in thir CSR vI (strand 2) to cyclodxtrin glu-canotransrass rom GH13_2 and in CSR vII (strand8) to -glucosidass rom GH13_31, rspctily [46].This rlatdnss as also sn in th olutionary tr oth ntir amily GH13 [6], hr subamilis GH13_1and GH13_2 ar adjacnt to ach othr, and th GH13_31

-glucosidass ar on th nighboring branch as a part oth so-calld oligo-1,6-glucosidas subamily clustr co-ring sral GH13 nzym spcicitis [67]. Structuraldtails can b ound in arious thr-dimnsional structurso th subamily GH13_2 cyclodxtrin glucanotransrass[86, 87] and subamily GH13_31 that contains oligo-1,6-glucosidass [88, 89], -glucosidas [90], dxtran glu-

cosidass [91, 92], and sucros isomrass [9395]. Thprsnc o a glutamic acid rsidu in th position i-3 romth transition-stat stabilizr, i.., th inariant aspartic acidrsidu at th strand 7 in CSR Iv (Fig. 2) may b con-sidrd as anothr charactristic squnc atur o mostsubamily GH13_1 mmbrs.

Subamilis GH13_5 and GH13_28

Th subamilis GH13_5 and GH13_28 includ bactrialliquying and sacchariying -amylass, rspctily. Tr-tiary structurs ar aailabl or thr typical rprsnta-

tis o th liquying -amylass o GH13_5, i.., romB. licheniormis (Fig. 1; [96, 97]), Bacillus stearothermo-philus [98] andB. amyloliqueaciens [99] and on sacchari-ying -amylas o GH13_28 rom Bacillus subtilis [100].Structurs ha bn dtrmind or additional GH13_5-amylass, .g., that rom Bacillus halmapalus [101], orhich also scondary (surac) binding sits r obsrd[102], to othrs rom Bacillus strains [103, 104], andn an xo-amylas rom Bacillus sp. 707 [105], i.., amaltohxaos-producing amylas [106] also classid insubamily GH13_5 (CAZy; [4]). Intrstingly, rom tholutionary point o i, th unclassid polyxtrmo-

philic -amylas AmyB rom Halothermothrix orenii [70]that posssss an N-trminal domain prcding th canoni-cal (/)8-barrl (Fig. 1), sms to b closly rlatd toGH13_5 subamily nzyms (Fig. 3).

Although th nzyms o ths to subamilis possssquit dirnt squncs, .g., on o th most obious di-rncs is th lngth and squnc o domain B [23], bothths -amylas subamilis blong to th so-calld amyl-as part in th olutionary tr o th ntir amily GH13coring subamilis 5, 6, 7, 15, 24, 27, 28, and 32 [6].

Latr, som ungal -amylass producd intracllu-larly r addd to th subamily GH13_5 [107]. Thsnzyms in som cass, .g., rom Histoplasma capsula-tum [108] and Paracoccidioides brasiliensis [109], smto b associatd ith irulnc by thir inolmnt inth production o an outr -(1,3)-glucan layr. Rcntly,this subamily as incrasd by addition o a potntial-amylass romArchaea (CAZy; [4]).

In subamily GH13_28, th prsnc o th SBD romamily CBM26 occurring as rpatd motis in -amylassrom lactobacilli [110, 111] is o a spcial intrst[112, 113]. It should b pointd out that th -amylass


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rom GH13_5 ar closly rlatd to plant and archaal-amylass rom subamilis GH13_6 and GH13_7(Fig. 3), hras thos rom GH13_28 ar closr to animal-amylass rom subamilis GH13_15 and GH13_24 [80,107, 114, 115].

Subamilis GH13_6 and GH13_7

Th subamilis GH13_6 and GH13_7 rprsnt mainly-amylass rom plants and Archaa, rspctily, hichr originally rald as constituting a group o closlyrlatd -amylass [114, 116]. Such a clos rlatdnss(Fig. 3) is rmarkabl not only du to a long taxonomicaldistanc btn prokaryotic archabactria and ukary-otic plants, but also bcaus o, or xampl, markd dir-ncs in thrmostability o archaal and plant -amylass.whil plant -amylass do not ha xcptional thrmo-stability, th GH13_7 -amylass ar, in gnral, producdby hyprthrmophilic archaons rom arious strains oto gnra, Pyrococcus and Thermococcus [117], andths nzyms tnd to b acti and stabl at tmpratursaround 80 C and highr.

Th bst charactrizd rprsntatis o th GH13_7subamily ar th -amylass rom Thermococcushydrothermalis [118], Thermococcus onnurineus [119],

Pyrococcus uriosus [120, 121], and Pyrococcus woesei[122], or hich also th thr-dimnsional structur hasalrady bn dtrmind [123]. Rcntly, a hypothti-cal -amylass romBacteria r assignd to th GH13_7subamily (CAZy; [4]) that as or a long priod consid-rd to b solly th archaal -amylas subamily [107,114, 115, 124]. All o ths hypothtical nzyms originat

intrstingly rom faobactria.On th othr hand, th subamily GH13_6 basically co-

rs highr plant -amylass and thos rom grn alga,but th bactrium Saccharophagus degradans as oundto contain in its gnom a plant-lik copy o -amylasin addition to a bactrial on, indicating a horizontal gntransr nt [115]. At prsnt, thr may b our GH13_6-amylass rom Bacteria, th on rom S. degradans,to rom strains oSpirochaeta thermophila and on romStigmatella aurantiaca (CAZy; [4]). with rgard to thbst charactrizd plant -amylass, ths ar th to bar-ly isozyms AMY1, a lo pI isozym [125] and AMY2,

a high pI isozym [126], ith sold thr-dimnsionalstructurs (Fig. 1) dtrmind both in r orms and ascomplxs [127130]. -Amylass rom othr highrplants, such as hat [131], ric [132], maiz [133], kid-ny ban [134], appl [135], Arabidopsis thaliana [136],banana [137], and othrs ar also includd in GH13_6(CAZy; [4]). Intrstingly, th plant -amylass blong toth GH13 -amylass haing th shortst polypptid chain(Fig. 2).

Surac binding sits r obsrd among th rpr-sntatis o both subamilis. In addition to our such sitssn in th structur o th archaal GH13_7 -amylas

rom P. woesei [123], th surac binding sits ha bnbst studid in to barly -amylas isozyms [129, 130,138140], spcially th on locatd in domain C o thlo pIisozym AMY1 calld a pair o sugar tongs [129]that shos no quialnt binding in th countrpart high pIisozym AMY2 [141].

Both GH13_6 and GH13_7 subamilis r originallyrald as to closly rlatd groups positiond in tholutionary tr on adjacnt branchs [114]. Mor rcntstudis ha indicatd [107, 115, 142, 143], hor, thatths to togthr ith th subamily GH13_5 blong toon olutionary clustr o-amylass (Fig. 3), both hnth phylogntic tr is basd on th alignmnt o CSRsonly and hn a substantial part o amino acid squncs,typically th catalytic domain, ar includd in th calcu-lation (.g., Fig. 2). This applis n in th olutionarytr o th ntir amily GH13 in th so-calld amylaspart [6]. Th subamilis GH13_6 and GH13_7 sharsral uniqu squnc aturs that discriminat thmrom th -amylass in th rmaining GH13 subamilis[114]. On o th most notabl aturs may b th Gly202(T. hydrothermalis-amylas numbring; [118]) at th nd

Fig. 2 Amino acid squnc alignmnt o GH13 -amylass rpr-snting th indiidual -amylas subamilis. Th alignd squncsspan th typical GH13 -amylass domain arrangmnt consisting ocatalytic (/)8-barrl ith domain B (insrtd btn th strand 3and hlix 3) and domain C (succding th TIM-barrl). Th namo an nzym is composd o th GH13 subamily numbr (xx orth rcntly dscribd -amylas rom Bacillus aquimaris and ??or -amylass rom A. haloplanktis, Bacteroides thetaiotaomicron,andHalothermothrix orenii AmyB that currntly ar still not assignd

any GH13 subamily in CAZy) ollod by th abbriation o thsourc (organism) and th UniProt accssion numbr. Not thr arto -amylass rom Halothermothrix orenii, th AmyA assignd tosubamily GH13_36 (UniProt Q8GPL8) and th AmyB not assigndas yt to any GH13 subamily (UniProt B2CCC1). Th organismsar abbriatd as ollos: Aspor, Aspergillus oryzae; Bacli,Bacilluslicheniormis; Horu, Hordeum vulgare; Pyco, Pyrococcus woesei;Tnmo, Tenebrio molitor; Sussc, Sus scroa (pancras); Arhy, Aer-omonas hydrophila; Bacsu, Bacillus subtilis; Stmli, Streptomyceslimosus; Hator, Halothermothrix orenii; Uncba, unculturd bact-rium; Bacaq, Bacillus aquimaris; Altha, A. haloplanktis; and Bctth,Bacteroides thetaiotaomicron. Th sn CSRs typical or th am-ily GH13 [46] and th catalytic triad ar highlightd in yellow andblue, rspctily. Th rsidus consrd inariantly ar markd byan asterisk blo th alignmnt. Th indiidual -amylas amilyGH13 domains ar indicatd as a colored lane above the alignment:blue catalytic domain A, green domain B, redC-trminal domain C.Th positions corrsponding to th dltd SBD o th amily CBM58in ??_Bctth_Q8A1G3 (SusG -amylas) ar signid by a magentalan. All squncs r rtrid rom th UniProt knoldg data-bas [251]. Th squnc alignmnts r don using th programClustal-w2 [252] and thn manually tund in ordr to maximizsqunc similaritis


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o th CSR II at th strand 4 o th catalytic (/)8-barrl(Fig. 2) th carbonyl group o hich srs as a spcic

ligand or th calcium ion in th complx structur o bar-ly -amylas (high pI isozym AMY2) ith acarbos[128] and could play th sam rol in th archaal countr-part, i.., th P. woesei-amylas [123].

Subamilis GH13_15, GH13_24 and GH13_32

Th -amylas subamilis GH13_15, GH13_24 andGH13_32 rprsnt mainly th -amylass rom inscts, ani-mals (including mammals) and Actinobactria, rspctily.Thir mutual rlatdnss (Fig. 3) as rald in 1994 [53]and conrmd in many subsqunt studis [107, 115, 142

144] including th comparison mad hn th ntir amilyGH13 as ocially diidd into subamilis [6].

A spcial cas is rprsntd by th -amylas rom thAntarctic psychrophilA. haloplanktis [145] that as dm-onstratd to xhibit clos similarity to animal -amylass[53, 115, 146], but up to no has not bn assignd to anyGH13 subamily (CAZy; [4]). Its thr-dimnsional struc-tur is aailabl [68, 69] and basd on structural studis[12] this -amylas as also ound to prorm a transgly-cosylation raction (Fig. 1). Togthr ith rlatd animal

-amylass (Fig. 3) it blongs to a group o th so-calldchlorid-actiatd -amylass [147, 148] and is a usul

modl or comparati studis [149].Among inscts, th -amylass blong to th subam-

ily GH13_15, and th -amylass rom a modl organ-ism Drosophila melanogaster [150] and rlatd ruit fisha bn th subjct o many olutionarily orintdstudis [151153]. Th trtiary structur is aailabl orth -amylas rom a yllo mal orm Tenebrio molitor[154, 155].

In th subamily GH13_24, coring animals rom, orxampl, molluscs [156, 157] and arthropods [158] to r-tbrats (i.., rogs, shs, birds, and mammals includinghuman bings; [159]), th bst knon rprsntatis ith

trtiary structurs aailabl ar pig pancratic -amylas[160163] plus thos rom human salia [164] and pan-cras [165]. Rcntly th structur o th -amylas o thsh Oryzias latipes as sold [166]. Th surac bindingsits r also ll rcognizd in trtiary structurs o s-ral subamily GH13_24 mmbrs, .g., in th -amylassrom pig pancras [163, 167169] and both human salia[13] and pancras [170].

Th third subamily GH13_32 intrstingly containsbactrial -amylass mostly rom actinomycts. Som o

(a) (b)

Fig. 3 eolutionary trs o GH13 -amylass rprsnting thindiidual -amylas subamilis. Th trs ar basd on th align-mnt o: a catalytic (/)8-barrl including domain B ith succd-ing domain C (653 positions; alignd in Fig. 2); and b sn GH13charactristic CSRs (52 rsidus; highlightd in Fig. 2). Th nams o

th -amylass ar xplaind in th lgnd to Fig. 2. Th olutionarytrs r calculatd as a Phylip-tr typ using th nighbor-joiningclustring [253] and th bootstrapping procdur [254] (th numbro bootstrap trials usd as 1,000) implmntd in th ClustalX pack-ag [252], and thn displayd ith th program Trvi [255]


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ths amylass may xhibit a maltotriohydrolas spcic-ity (eC 3.2.1.116), .g., thos rom Thermobifda usca[171] and Brachybacterium sp. [172], both blonging toActinobactria. Such -amylass rom Actinobactria arotn calld animal-typ -amylass [115]. In this sub-amily, th halophilic -amylas rom Kocuria varians hasbn dscribd rcntly [173] as possssing, C-trminal to

th catalytic domain, to tandm copis o an SBD o thamily CBM25 [174, 175]. A similar domain arrangmntas ound priously in th -amylas rom Bacillus sp.No. 195 [176] that also blongs to th subamily GH13_32(CAZy; [4]). Typical GH13_32 -amylass rom strpto-mycts [177, 178] may also contain an SBD, usually oth amily CBM20 [41] at thir C-trminus. A ry rcntolutionary study has idntid th subamily GH13_32typ o-amylas in basidiomycts o ungi, i..,Eucarya[179] and th authors, in agrmnt ith anothr study[180], concludd that th gn donor may ha originatdrom Actinobactria.

Subamily GH13_27

Th subamily GH13_27 cors a small group (~50squncs) o bactrial -amylass (Fig. 3) rcognizd ashomologous bor th CAZy classication as stab-lishd [181]. In most studis [53, 80, 115] this subamilyis usually rprsntd by to xprimntally charactrizd-amylass rom Aeromonas hydrophila [182] and Xan-thomonas campestris [181]. Although no thr-dimnsionalstructur is aailabl or any GH13_27 -amylas, thr isno doubtbasd on squnc comparisonths nzyms

ar typical 3-domain GH13 -amylass (i.., ithout addi-tional domains such as SBDs). In addition to th to rp-rsntatis rom A. hydrophila and X. campestris, to-amylass rom Pseudomonas sp. KFCC 10818 ha bnclond, xprssd, squncd, and charactrizd [183, 184].

Subamily GH13_36

Th subamily GH13_36 sms to contain a group o-amylass possssing rlatd GH13 nzym spcicitis[185]. This subamily as originally dnd as th groupo amylolytic nzyms ith an intrmdiary position inth olutionary tr btn th polyspcic subamiliso oligo-1,6-glucosidas and nopullulanas [67]. That pro-posal as mainly basd on a spcic squnc in th CSRvtypically QPDLN and MPKLN or oligo-1,6-glucosi-dass and nopullulanass, rspctily, th intrmdiarygroup bing charactrizd by MPDLN (Fig. 2). Noadays,th oligo-1,6-glucosidas and nopullulanas subami-lis cor sral CAZy-curatd GH13 subamilis [6].Th intrmdiary group as, hor, assignd to thsubamily GH13_36 only rcntly (CAZy; [4]). Only 11

GH13_36 mmbrs ha bn biochmically charactrizdto any xtnt, although in som cass, ths nzyms rdscribd just as an amylas [185], .g., th nzymsrom Dictyoglomus thermophilum AmyC [186] and Bacil-lus megaterium [187]. Th actiity toards -1,6-branchdglucans togthr ith a transras ability r, hor,dmonstratd or th amylolytic nzym rom B. mega-

terium [188, 189] as ll as or th priplasmic on romX. campestris [190]. Furthrmor, th GH13_36 amyl-as rom Paenibacillus polymyxa as abl to rlas pan-os rom pullulan [191, 192], th on romBacillus clarkiixhibitd prdominating actiity toard -cyclodxtrins[193] and th lipoprotin amylas rom Anaerobrancagottschalkii shod both cyclodxtrin-hydrolyzing andtransglycosylating actiitis [194]. No additional acti-ity as obsrd or th othr lipoprotin -amylas romThermotoga maritima [195] or or th halothrmophilic-amylas romH. orenii [196], or hich th thr-dimn-sional structur is aailabl as th only rprsntati o th

subamily GH13_36 [197].

Rcntly stablishd subamily GH13_37

Th subamily GH13_37 is th most rcntly stablishd-amylas subamily in CAZy (CAZy; [4]). It as cratdbasd on isolation and phylogntic analysis o a nol-amylas dsignatd AmyP rom a marin mtagnomiclibrary [198]. Th nzym as latr shon to xhibit thability to dgrad ra starch [199]. Currntly this subamilycontains, in addition to th AmyP,


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closly rlatd to thos rom subamily GH13_1, spciallyi only th GH13 charactristic CSRs IvII ar considrd(Fig. 3). Th -amylass rom GH13_37, togthr ith thabo-mntiond GH13_36 intrmdiary -amylass andth GH13_19 maltohxaos-producing amylass, round to occupy th adjacnt clustr in th olutionarytr [71]. Th xclusi squnc atur o this nly pro-

posd GH13 -amylas subamily is th prsnc o toconscuti tryptophans (Trp201 and Trp202, B. aquima-ris-amylas numbring), locatd at hlix 3 (Fig. 2) pr-cding strand 4 (i.., th CSR II) o th catalytic (/)8-barrl domain, that may indicat a surac binding sit[71]. Th sam atur is prsnt in -amylas homologusrom Geobacillus thermoleovorans [202] and Anoxybacil-lus sp. SK34 and sp. DT31 [203]. Th rcntly soldthr-dimnsional structur o th G. thermoleovorans-amylas has shon [204] that th abo-mntiondtryptophan pair is positiond in a rgion ith additionalaromatic rsidus xhibiting a structural rsmblanc to

th nopullulanas subamily GH13_20, spcially to thcyclomaltodxtrinas rom Flavobacterium sp. 92 [205]. Inth lattr nzym th aromatic rgion intracts ith domainN, hich is not prsnt in -amylass. Moror, th sc-ond tryptophan o th xclusi tryptophan pair (w205, G.thermoleovorans-amylas numbring) is not xposd toth solnt but burid [204]. Furthr, it as pointd out thatth G. thermoleovorans-amylas rsmbls in structur,not only GH13_1 and GH13_20 nzyms but also somrom th GH13_2 subamily [204] that contains cyclodx-trin glucanotransrass [6].

Notably thr is anothr intrsting and yt unclassi-

d -amylas ith thr-dimnsional structur alradysoldth SusG protin rom Bacteroides thetaio-taomicron [38]. It is rlatd to subamilis GH13_36 andGH13_37 and, in a idr sns also to th currntly unclas-sid group rprsntd by marinB. aquimaris-amylasmntiond abo (Fig. 3). Th -amylas SusG is ncoddby a mmbr o th sus (starch utilization systm) locus oth human gut symbiont B. thetaiotaomicron [206208]and rprsnts th only GH13 -amylas ith an SBDinsrtd in domain B (Fig. 1); in this particular cas thSBD is o amily CBM58 [38].

Family GH57

A n amily, GH57, containing -amylass as cr-atd in 1996 [3] atr long orts to nd th amily GH13squnc aturs in to supposd -amylass, on romth thrmophilic bactrium D. thermophilum [209] andth othr rom th hyprthrmophilic archaon P. uriosus[210]. en atr th amily GH57 as stablishd, thpossibility o a rlationship btn th to amilis GH13

and GH57 as not dnitily liminatd, although it hadprod impossibl to align conincingly th to abo-mntiond GH57 nzyms ith rprsntatis o GH13-amylass [211]. Th rst GH57 crystal structur, ho-r, sold or th 4--glucanotransras rom Thermo-coccus litoralis in 2003, unambiguously conrmd that thGH57 amily cannot blong to th GH-H clan [212].

Noadays th amily GH57 rprsnts a scond andsmallr -amylas amily [213]. It contains ~900 mmbrs

Fig. 4 a eolutionary tr shoing th -amylas rprsntatio th amily GH57 among th othr GH57 nzym spcicitistogthr ith th -amylas rprsntati o th amily GH119. Thtr is basd on th alignmnt o th GH57 charactristic CSRs(36 rsidus). Th nam o an nzym is composd o th GH57 orGH119 amily numbr ollod by th abbriation o spcic-ity (in capitals), abbriation o sourc (organism) and th UniProtaccssion numbr. All squncs r rtrid rom th UniProtknoldg databas [251]. Th spcicitis ar abbriatd as ol-

los: AAMY -amylas, PAMY putati -amylas-lik protin,MGA maltognic amylas, AMY unspcid amylas, APU amy-lopullulanas, APU-CMD amylopullulanas-cyclomaltodxtrinas,BE branching nzym, BE-AAMY -amylas-branching nzym,4AGT 4--glucanotransras, AGAL -galactosidas. Th organ-isms ar abbriatd as ollos: Bacci, Bacillus circulans; Bccth,Bacteroides thetaiotaomicron; Mccja, Methanocaldococcus jan-naschii; Pycu, Pyrococcus uriosus; Sttma, Staphylothermus mari-nus; Thchy, Thermococcus hydrothermalis; Thcko, Thermococcuskodakaraensis; Thcli, Thermococcus litoralis; Thtma, Thermotogamaritima; Uncba, unculturd bactrium. Th olutionary tr ascalculatd as a Phylip-tr typ using th nighbor-joining clustr-ing [253] and th bootstrapping procdur [254] (th numbr obootstrap trials usd as 1,000) implmntd in th ClustalX pack-ag [252], and thn displayd ith th program Trvi [255]. bStructural modls o -amylass rom amilis GH57 and GH119.Let suprimposd modld structurs o th GH57 Methanocaldo-coccus jannaschii -amylas (blue) and GH119 Bacillus circulans-amylas (magenta). Th -hlical bundl succding th catalytic(/)7-barrl as modld only in th GH57 -amylas. Th rectan-gle indicats a dtaild i on th right. Righta clos-up ocusdon prdictd catalytic rsidus o th -amylass rom GH57 (Glu145and Asp237) and GH119 (Glu231 and Asp373). Both modls rsuprimposd ith th ral structur o GH57 Thermococcus lito-ralis 4--glucanotransras (PDB cod: 1K1Y; [212]; not shon).Th structural modls o -amylass r obtaind rom th Phyrsrr [245] basd on th GH57 4--glucanotransras tmplat asollos: rsidus Mt1-Tyr356 o th GH57 -amylas [216] andThr121-Asp429 o th GH119 -amylas [242]. Th suprimposd

part cors 218 C-atoms ith a 0.75 root-man squar diation;th suprimposition as don using th MultiProt srr [256]. Acar-bos-occupying subsits 1 through +3 [257] rom th complx ithGH57 4--glucanotransras structur [212] is shon. Th struc-turs r isualizd ith th program wbLabvirLit (Molcu-lar Simulations, Inc.). c Squnc logos o-amylass rom amilisGH57 rom Archaa and Bactria and GH119. CSR-1, rsidus 15;CSR-2, rsidus 611; CSR-3, rsidus 1217; CSR-4, rsidus 1827; CSR-5, rsidus 2836.Asterisks signiy th catalytic nuclophil(glutamic acid) in position 15 (in CSR-3) and proton donor (asparticacid) in position 20 (in CSR-4). Th logos ar basd on idntiyingth CSRs in both amilis [217, 242] as ollos: or 59 -amylassquncs (47 rom Archaa and nin rom Bactria) rom amilyGH57 and or six bactrial GH119 -amylass. Squnc logos rcratd using th wbLogo 3.0 srr [258]


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all originating rom prokaryots ith th approximatArchaa:Bactria ratio 3:1 and, lik th main -amylasamily GH13, th amily GH57 is a polyspcic amily(Fig. 4a; CAZy; [4]). Intrstingly, th to undamn-tal amily mmbrs, hich r originally considrdto b -amylass [209, 210], no ar both rcognizd as

4--glucanotransrass: th nzym rom D. thermo-philum as r-aluatd as a glucanotransras in 2004[214], hras a transras actiity or th on romP. uriosus as idntid immdiatly [215]. Thr ar ll-stablishd nzym spcicitis in th amily GH57[216, 217], i.., -amylas [218], 4--glucanotransras

(a)

(b)

(c)


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[209, 210, 219221], amylopullulanas [222225], branch-ing nzym [226228], and n -galactosidas [229].Noticably, to additional amylolytic spcicitis in thamily GH57 may b dnd by to partially charactr-izd GH57 mmbrs: th PF0870 opn rading ram romth P. uriosus gnom [230] and a non-spcid amylaso an unculturd bactrium isolatd rom a hydrothrmal

nt in th Atlantic Ocan [231]. Rcntly, a GH57 nzymas dscribd possssing dual spcicity: th amylopul-lulanas rom th archaon Staphylothermus marinus alsoxhibits cyclomaltodxtrinas actiity [232]. Dual spcic-ity may also b ound or th AmyC nzym rom T. mar-itima, dscribd originally as an -amylas [226], but itssqunc-structural aturs clarly indicat it may haalso branching nzym actiity [217].

Although amily GH57 shars th rtaining ractionmchanism ith th main -amylas amily GH13 [87,228], a (/)7-barrl (i.., an incomplt TIM barrl) isadoptd as th old baring th catalytic rsidus [212, 228,

233235]. Any GH57 mmbr can b charactrizd by CSRsidntid originally by Zona t al. [236] and rndrcntly by Blsak and Janck [217]that ar clarly di-rnt rom thos charactristic or amily GH13 [46]. Thcatalytic machinry also discriminats th amilis GH57and GH13 rom ach othr, bcaus a glutamic acid atstrand 4 (Glu123 in T. litoralis 4--glucanotransras)and aspartic acid at strand 7 (Asp214) o th (/)7-barrlact as th catalytic nuclophil and proton donor, rspc-tily [212, 228]. Rmarkably, in GH57 nzyms, thr isno third catalytic amino acid rsidu lik th transition-stat stabilizr ncssary in amily GH13 [10, 87, 237]. Th

(/)7-barrl domain is succdd in ry GH57 mmbrby a hlical sgmnt consisting o a bundl o 34 -hlics[216, 217]. Sinc th CSR-5 positiond in this -hlicalbundl as rald to contain unctionally ssntial rsi-dus [228], it as proposd that both th barrl and th hl-ical bundl togthr orm th catalytic ara o nzyms inth amily GH57 [217].

-Amylas and its putati homologus

It should b pointd out that although th amily GH57 isconsidrd to b an -amylas amily, th tru -amylas(eC 3.2.1.1) nzym spcicity has still not bn conrmdunambiguously. Th only idnc coms rom th studyby Kim t al. [218] ho shod that th -amylas romth mthanognic archaon Methanococcus jannaschiidgrads solubl starch. Th uncrtainty concrning thxact spcicity has arisn rom th act th -amylas asalso abl to dgrad pullulan at a rlati rat o 82 % othat dtrmind or starch [218, 238]. Thus th potntial-amylas, idntid rst as a hypothtical MJ1611 opnrading ram in th M. jannaschii gnom [239], is th

lading amily GH57 mmbr arranting th -amylasdsignation [240].

what is, hor, mor intrsting is th obsrationthat amily GH57 contains mmbrs, i.., th so-calld-amylas-lik protins, xhibiting clar squnc aturso th -amylas rom M. jannaschii, but simultanouslylacking on or both catalytic rsidus [216]. with rgard to

thir origin, GH57 -amylass com prdominantly romArchaa (~80 %), hras th GH57 -amylas-lik pro-tins originat mostly rom Bactria (~85 %). Both o thsgroups contain protins approximatly 500 amino acid rsi-dus long that xhibit a high dgr o squnc idntity toach othr, spcially in th CSRs that ha bn suggstdto b squnc ngrprints o th indiidual amily GH57nzym spcicitis [217]. Th most substantial dirncdiscriminating th -amylass and -amylas-lik protinso amily GH57 rom ach othr is th prsnc o bothcharactristic catalytic rsidus in -amylass (Fig. 4b)and th lack o on or both o ths rsidus in thir homo-

logus [216]. Th catalytic nuclophil locatd at th strand4 o th catalytic (/)7-barrl in CSR-3 (Glu145 in M.

jannaschii -amylas) and proton donor at th strand 7in CSR-4 (Asp237) ar most rquntly substitutd by asrin and glutamic acid, rspctily, in amylas-lik pro-tins that ar ound mainly in to gnra Bacteroides andPrevotella, both rom th ordr Bactroidals o Bactria.

Th amily GH57 nzyms ith spcicitis o amylop-ullulanas, branching nzym and 4--glucanotransrascontain som squnc-structural sgmnts at thir C-tr-mini additional to catalytic (/)7-barrl ith succd-ing thr-hlix bundl [212, 228, 234, 235], hil both-amylass and -amylas-lik protins sm to b com-posd o only th barrl and hlical bundl [217]. A similarto-domain arrangmnt is also ound in -galactosidass,but th domain coring th -hlical rgion in -amylasssms usually to b ~50100 rsidus longr than that pr-snt in th -galactosidass [217]. with rgard to th N-tr-minus, thr is probably no signal pptid in -amylassand -amylas-lik protins sinc th CSR-1 is almostxclusily locatd ry clos to th N-trminus [216] andno inormation is aailabl on a signal pptid or th onlycharactrizd -amylas romM. jannaschii [218].

Th uniqu squnc aturs that discriminat both-amylass and -amylas-lik protins rom th rmain-ing amily GH57 nzym spcicitis as ll as rom achothr ar sn in thir CSRs and bst rprsntd by thirsqunc logos. Thy r suggstd to dn th so-calld squnc ngrprints o a gin nzym spcicity[217]. Basd on a dtaild bioinormatics analysis o 367squncs rsulting in crating th squnc logos or GH57 nzym spcicitis, th positions 1, 12, 13, 21, 27,and 3536 r rald to b spcically charactristicor -amylas as ollos (Fig. 4c): (1) positions 1 (CSR-1)


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and 12 (CSR-3) contain mostly glutamic acid or glutaminand arginin or glutamic acid, rspctily, ith all ourrmaining ll-stablishd GH57 spcicitis bing char-actrizd by a histidin and tryptophan in th corrspond-ing positions; (2) positions 13 (CSR-3) and 21 (CSR-4) arxclusily occupid by inariant asparagin and tyrosin,rspctily, th othr spcicitis possssing dirnt

rsidus thr and not so strictly consrd; (3) th posi-tion 27 (CSR-4) is an inariant histidin, but this positionis not so uniqu or -amylass sinc a corrspondinghistidin can also b ound in amylopullulanass; and (4)positions 3536 at th nd o logo (CSR-5) consist o toadjacnt tyrosins and may rprsnt th most unambigu-ous GH57 -amylas atur bcaus a tyrosin rsidu hasnot bn sn at ths positions in any GH57 squncsascribd to a spcicity othr than that o -amylas. Itshould b mphasizd hr that ths last to positions ina GH57 squnc logo (positions 3536; CSR-5) rprsnta squnc ngrprint that most rliably distinguishs th

indiidual nzym spcicitis rom ach othr [217].Squnc logos ral also spcic dirncs btn

-amylass and thir -amylas-lik protin countrparts[216]. For xampl: (1) th inariant prolin in position 5(th nd o CSR-1) in -amylass is otn substitutd by anisolucin in bactrial -amylas-lik protins; (2) th posi-tion 7 (CSR-2) is rquntly occupid by a cystin i ithro th to, i.., an -amylas and/or an -amylas-lik pro-tin, originats rom Bacteria (or th -amylas-lik pro-tins rom th gnus Bacteroides, thr ar to adjacntcystins in th CSR-2); (3) highly spcic or -amylassis th prsnc o thr aromatic rsidus in positions 18,

21 and 24 (CSR-4), hras only th position 18 mayb considrd as aromatic on in th -amylas-lik pro-tins; and (4) th last thr positions (3436; CSR-5) oth -amylas squnc logo ar mostly aromatic rsidus,too, th middl position 35 bing rplacd by an argininin th -amylas-lik protins. Ths spcic dirncsbtn th nzymatically acti -amylass and thirmost probably inacti -amylas-lik countrparts romth amily GH57 ar, hor, still smallr than th di-rncs btn th -amylass and th othr ll-stab-lishd nzyms spcicitis ithin GH57, as indicatd byth olutionary tr (Fig. 4a).

Family GH119

Th amily GH119 as stablishd in 2006 olloingth study by watanab t al. [241] ho dscribd a nol-amylas as a product o th gnIgtZromBacillus circu-lans AM7 and ound no obious squnc similarity to any-amylas rom ithr amily GH13 or GH57. Th spci-city o-amylas as ascribd to th IgtZ protin basd

on production o glucos and maltooligosaccharids up tomaltopntaos rom maltooligosaccharids longr thanour glucos units, amylos and solubl starch [241]. Th-amylas IgtZ also contains SBDs o amilis CBM20 (1copy) and CBM25 (2 copis). Th amily GH119 may thusb considrd as th third CAZy -amylas amily, but itis orth mntioning that currntly, i.., 7 yars atr it as

cratd in th CAZy databas, th amily is still ry smallsinc only hypothtical bactrial protins obtaindrom gnom squncing projcts ha bn addd to th-amylas IgtZ dscribd originally (CAZy; [4]). Untilrcntly, hn a clos rlatdnss o th amily GH119 toGH57 as rald (Fig. 4b; [242]), inormation on amilyGH119 concrning squnc-structural dtails (.g., cata-lytic machinry and old) as, in act, lacking [243].

A rlatdnss to th amily GH57

Basd on a dtaild in silico analysis that inold com-

parison o amino acid squncs o all amily GH119mmbrs (CAZy; [4]) in combination ith th BLAST tool[244] and trtiary structur modling at th Phyr-2 [245]and SissModl [246] srrs, an unambiguous olu-tionary rlatdnss as rald btn amilis GH119and GH57 [242]. Th CSRs charactristic o th am-ily GH57 [217, 236] r idntid in th squncs oall six GH119 mmbrs. It as thus possibl to prdict thGH119 catalytic rsidus, i.., Glu231 and Asp373 in thsqunc o th -amylas IgtZ rom B. circulans, hichar consrd inariantly in GH119 (Fig. 4c). This prdic-tion is urthr supportd by thr-dimnsional structur

modling indicating that th amily GH119 shars ith thamily GH57 both th catalytic (/)7-barrl old and cat-alytic machinry (a glutamic acid as th catalytic nuclo-phil and an aspartic acid as th proton donor at th strands4 and 7 o th barrl; Fig. 4b). Dspit th clar similar-ity, th amily GH119 rtains its on idntity in th olu-tionary tr common or both amilis (Fig. 4a) rfctingits charactristic squnc aturs also in th CSRs [242].As suggstd, th cration o a nol CAZy GH clan oamilis GH57 and GH119 is thus highly probabl but thisnds to b conrmd by soling th trtiary structur andidntiying xprimntally th catalytic rsidus in a amilyGH119 rprsntati [242].

Family GH126

Th last CAZy GH amily supposdly containing th spci-city o -amylas is th amily GH126, cratd on thbasis o a rport by Ficko-Blan t al. [17]. Thy assssdth CPF_2247 protin product rom th Clostridium per-

ringens ATCC 13124 gnom as an -amylas sinc th


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nzym shod actiity on maltooligosaccharids, amyl-

os and glycogn, but no attmpt as mad to sho thatanomric conguration is consrd. Rmarkably, althoughth CPF_2247 dns its on amily GH126, hich cur-rntly also contains approximatly 60 hypothtical protinsrom Bactria (CAZy; [4]), it xhibits ~20 % squncidntity to ndo--1,4-glucanass rom th amily GH8[17]. Moror, th CPF_2247 is structurally most closlyrlatd to GH8 ndoglucanas ClA [247] and GH48 cllo-biohydrolas ClS [248] both rom Clostridium thermocel-lum. Ths to amilis orm togthr th clan GH-M(CAZy; [4]). Th nzyms rom th clan GH-M mploy,hor, an inrting mchanism o glucosidic bond cla-ag and th CPF_2247 -amylas sms to shar iththm, in addition to a catalytic (/)6-barrl old, its cata-lytic nuclophil (Glu84) plus a othr inariantly con-srd and unctionally important rsidus (Fig. 5; [17]).Ths acts ar not consistnt ith th gnral dnition o-amylas nzym spcicity [9, 87] mploying th rtain-ing mchanism, that has bn conrmd by crystal struc-turs o arious -amylass rom amily GH13 [11, 12, 14,73, 100, 101, 123, 128, 130, 148, 162] and rprsntatisrom amily GH57 [212, 228].

Dspit ths apparnt inconsistncis, monitoring

th dpolymrization o arious polysaccharids by thCPF_2247 using thin-layr chromatography rald claractiity on amylos and glycogn (ith production o amixtur o maltooligosaccharids rom maltos to malto-hptaos), but no actiity on pullulan and cllulos [17].Furthrmor, raction products rom th sris o malto-trios to maltohptaos tstd by high-prormanc anionxchang chromatography shod that maltopntaos isth minimal oligosaccharid substrat or CPF_2247, romhich th nzym rmos a singl glucos rsidu [17].This mans that, as ll as th diculty o undrstandingho an -amylas could shar th catalytic mchanismith inrting -glucanass, thr is also som uncr-tainty concrning th ndo/xo ashion o action o thisrmarkabl GH126 amylolytic nzym CPF_2247 rom C.

perringens.

Conclusions

As documntd in th prsnt ri, -amylas ranksamong th most rquntly occurring CAZy. It is likly

(a) (b)

Fig. 5 Structur o th -amylas o amily GH126. a GH126structur o th -amylas rom Clostridium perringens (PDB cod:3ReN; [17]) shoing th (/)6-barrl ith highlightd rsidus(colord by lmnt) inold in catalysis: Glu84 (gnral acid),Asp136 (gnral bas), and Tyr194 (contributing to catalysis). bSuprimposition oClostridium perringens GH126 -amylas (blue)ith Clostridium thermocellum GH8 ndoglucanas ClA (redPDBcod: 1KwF; [247]). Th suprimposd part cors 193 C-atomsith a 1.93 root-man squar diation; th suprimposition asdon using th MultiProt srr [256]. Th dtaild i ocuss onth catalytic rsidus in th structur o GH8 ndoglucanas ClA

(Glu95 and Asp278 ith Tyr215) and th proposd rsidus in thGH126 -amylas (Glu84 and Asp136 ith Tyr194). Cllopnta-os occupying subsits 3 through +2 [257] in complx ith GH8ndoglucanas ClA is shon. A comparison ith th amily GH15glucoamylas, i.., an -glucan-acti nzym ith (/)6-barrlcatalytic old mploying th inrting mchanism [259], rals lsssimilarity sinc th suprimposd part btn th GH126 -amylasand Aspergillus niger GH15 glucoamylas [260] cord only 146C-atoms ith a 1.97 root-man squar diation. Th structursr rtrid rom th PDB [250] and isualizd ith th programwbLabvirLit (Molcular Simulations, Inc.)


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that -amylass ar prsnt in amilis GH57 and GH119,and possibly n in GH126, but -amylas is considrdth main rprsntati o amily GH13knon gnr-ally as th main -amylas amily. This GH amily ormsa clan togthr ith amilis GH70 and GH77, but in thlattr to, no -amylas spcicity has bn ound. withinth amily GH13, hor, th -amylass dn sral

GH13 subamilis, th mmbrs o hich xhibit closrsqunc-structural similaritis than obsrd or th am-ily as a hol. Nrthlss, dspit mor than 13,500squncs classid currntly in th amily GH13, all thmmbrs shar 47 CSRs, th -amylas-typ o (/)8-barrl catalytic domain, catalytic machinry, and rtainingraction mchanism. Th -amylas classid in amilyGH57 mploys th sam rtaining mchanism as GH13-amylas. All th amily GH57 mmbrs dir, hor,rom thos o amily GH13 in that thy possss thir on CSRs and adopt a (/)7-barrl old (i.., an incompltTIM-barrl) baring catalytic machinry dirnt rom that

usd in th amily GH13. All o ths GH57 charactristicsar likly to b shard by amily GH119, hich is th thirdGH amily containing -amylas spcicity. Th last GHamily that has bn indicatd to contain th spcicity oan -amylas, i.., th amily GH126, is o spcial intrstand th situation may b clarid hn urthr biochmi-cal charactrization and structur dtrmination ha bncarrid out, sinc protins o th amily GH126 xhibit anunambiguous homology to -glucan-acti hydrolass oamilis GH8 and GH48 that mploy an inrting ractionmchanism.

Futur discoris rlating to -amylass may sho

idr, but rlatd, spcicitis and n structurs, gi-ing n amilis ithin th CAZy systm, but thoroughbiochmical charactrization and structur dtrminationis rquird bor n and surprising conclusions can baccptd.

Acknowledgments SJ thanks th Sloak Rsarch and Dlop-mnt Agncy or nancial support undr contract No. LPP-0417-09and th Sloak Grant Agncy veGA or Grant No. 2/0148/11. BSthanks th Danish Rsarch Council or Indpndnt Rsarch | Natu-ral Scincs (FNU) or nancial support undr Grant No. 09-072151.

Reerences

1. Hnrissat B (1991) A classication o glycosyl hydrolass basdon amino acid squnc similaritis. Biochm J 280:309316

2. Hnrissat B, Bairoch A (1993) N amilis in th classica-tion o glycosyl hydrolass basd on amino acid squnc simi-laritis. Biochm J 293:781788

3. Hnrissat B, Bairoch A (1996) Updating th squnc-basd classication o glycosyl hydrolass. Biochm J 316:695696

4. Cantarl BL, Coutinho PM, Rancurl C, Brnard T, Lombardv, Hnrissat B (2009) Th Carbohydrat-Acti enZyms

databas (CAZy): an xprt rsourc or glycognomics.Nuclic Acids Rs 37:D233D238

5. Hnrissat B, Dais GJ (1997) Structural and squnc-basdclassication o glycosid hydrolass. Curr Opin Struct Biol7:637644

6. Stam MR, Danchin eG, Rancurl C, Coutinho PM, Hnris-sat B (2006) Diiding th larg glycosid hydrolas amily 13into subamilis: toards improd unctional annotations o-amylas-rlatd protins. Protin eng Ds Sl 19:555562

7. Aspborg H, Coutinho PM, wang Y, Brumr H 3rd, HnrissatB (2012) eolution, substrat spcicity and subamily classi-cation o glycosid hydrolas amily 5 (GH5). BMC eol Biol12:186

8. St John FJ, Gonzalz JM, Pozharski e (2010) Consolidationo glycosyl hydrolas amily 30: a dual domain 4/7 hydrolasamily consisting o to structurally distinct groups. FeBS Ltt584:44354441

9. MacGrgor eA (1988) -Amylas structur and actiity. J Pro-tin Chm 7:399415

10. MacGrgor eA, Janck S, Snsson B (2001) Rlationship osqunc and structur to spcicity in th -amylas amily onzyms. Biochim Biophys Acta 1546:120

11. Brzozoski AM, Dais GJ (1997) Structur o thAspergillusoryzae-amylas complxd ith th inhibitor acarbos at 2.0 rsolution. Biochmistry 36:1083710845

12. Aghajari N, Roth M, Hasr R (2002) Crystallographic idnco a transglycosylation raction: trnary complxs o a psy-chrophilic -amylas. Biochmistry 41:42734280

13. Ramasubbu N, Ragunath C, Mishra PJ (2003) Probing th rolo a mobil loop in substrat binding and nzym actiity ohuman saliary amylas. J Mol Biol 325:10611076

14. Li C, Bgum A, Numao S, Park KH, withrs SG, Brayr GD(2005) Acarbos rarrangmnt mchanism implid by thkintic and structural analysis o human pancratic -amylasin complx ith analogus and thir longatd countrparts.Biochmistry 44:33473357

15. Brzozoski AM, Lason DM, Turknburg JP, Bisgaard-Frantzn H, Sndsn A, Borchrt Tv, Dautr Z, wilson KS,

Dais GJ (2000) Structural analysis o a chimric bactrial-amylas. High-rsolution analysis o nati and ligand com-plxs. Biochmistry 39:90999107

16. Janck S, Snsson B, MacGrgor eA (1995) Charactris-tic dirncs in th primary structur allo discrimination ocyclodxtrin glucanotransrass rom -amylass. Biochm J305:685686

17. Ficko-Blan e, Stuart CP, Boraston AB (2011) Structural analy-sis o CPF_2247, a nol -amylas rom Clostridium perrin-gens. Protins 79:27712777

18. Snsson B (1988) Rgional distant squnc homologybtn amylass, -glucosidass and transglucanosylass.FeBS Ltt 230:7276

19. Kuriki T, Imanaka T (1989) Nuclotid squnc o th nopul-lulanas gn rom Bacillus stearothermophilus. J Gn Micro-

biol 135:1521152820. MacGrgor eA, Snsson B (1989) A supr-scondary struc-tur prdictd to b common to sral -1,4-d-glucan-claingnzyms. Biochm J 259:145152

21. Jsprsn HM, MacGrgor eA, Sirks MR, Snsson B (1991)Comparison o th domain-ll organization o starch hydro-lass and rlatd nzyms. Biochm J 280:5155

22. Snsson B, Janck S (2013) Glycosid hydrolas amily 13.CAZypdia. http://.cazypdia.org/. Accssd 18 Mar 2013

23. Janck S, Snsson B, Hnrissat B (1997) Domain olutionin th -amylas amily. J Mol eol 45:322331

24. Fort J, d la Ballina LR, Burghardt He, Frrr-Costa C, TurnayJ, Frrr-Orta C, Uson I, Zorzano A, Frnandz-Rcio J, Orozco
http://www.cazypedia.org/http://www.cazypedia.org/


16/22

. Janeek et al.

1 3

M, Lizarb MA, Fita I, Palacin M (2007) Th structur ohuman 4F2hc ctodomain proids a modl or homodimriza-tion and lctrostatic intraction ith plasma mmbran. J BiolChm 282:3144431452

25. Gabrisko M, Janck S (2009) Looking or th ancstry o thhay-chain subunits o htromric amino acid transportrsrBAT and 4F2hc ithin th GH13 -amylas amily. FeBS J276:72657278

26. MacGrgor eA (1993) Rlationships btn structur

and actiity in th -amylas amily o starch-mtabolisingnzyms. Starch/Stark 45:232237

27. Janck S (1994) Paralll /-barrls o-amylas, cyclodx-trin glycosyltransras and oligo-1,6-glucosidas rsus thbarrl o -amylas: olutionary distanc is a rfction ounrlatd squncs. FeBS Ltt 353:119123

28. Snsson B (1994) Protin nginring in th -amylas am-ily: catalytic mchanism, substrat spcicity, and stability.Plant Mol Biol 25:141157

29. Kuriki T, Imanaka T (1999) Th concpt o th -amylas am-ily: structural similarity and common catalytic mchanism. JBiosci Biong 87:557565

30. an dr Maarl MJ, an dr vn B, Uitdhaag JC, LmhuisH, Dijkhuizn L (2002) Proprtis and applications o starch-conrting nzyms o th -amylas amily. J Biotchnol94:137155

31. Matsuura Y, Kusunoki M, Harada w, Kakudo M (1984) Struc-tur and possibl catalytic rsidus o Taka-amylas A. J Bio-chm 95:697702

32. Bannr Dw, Bloomr AC, Ptsko GA, Phillips DC, PogsonCI, wilson IA, Corran PH, Furth AJ, Milman JD, Oord Re,Priddl JD, waly SG (1975) Structur o chickn muscl tri-os phosphat isomras dtrmind crystallographically at 2.5angstrom rsolution using amino acid squnc data. Natur255:609614

33. Farbr GK, Ptsko GA (1990) Th olution o / barrlnzyms. Trnds Biochm Sci 15:228234

34. Pnninga D, an dr vn BA, Kngtl RM, an Hijum SA,Rozboom HJ, Kalk KH, Dijkstra Bw, Dijkhuizn L (1996)

Th ra starch binding domain o cyclodxtrin glycosyl-transras rom Bacillus circulans strain 251. J Biol Chm271:3277732784

35. Ab A, Tonozuka T, Sakano Y, Kamitori S (2004) Complxstructurs o Thermoactinomyces vulgaris R-47 -amylas 1ith malto-oligosaccharids dmonstrat th rol o domain Nacting as a starch-binding domain. J Mol Biol 335:811822

36. Boraston AB, Haly M, Klassn J, Ficko-Blan e, Lammrtsan Burn A, La v (2006) A structural and unctional analy-sis o-glucan rcognition by amily 25 and 26 carbohydrat-binding moduls rals a consrd mod o starch rcogni-tion. J Biol Chm 281:587598

37. an Burn AL, Boraston AB (2007) Th structural basis o-glucan rcognition by a amily 41 carbohydrat-binding mod-ul rom Thermotoga maritima. J Mol Biol 365:555560

38. Koropatkin NM, Smith TJ (2010) SusG: a uniqu cll-mm-bran-associatd -amylas rom a prominnt human gut sym-biont targts complx starch molculs. Structur 18:200215

39. Snsson B, Jsprsn H, Sirks MR, MacGrgor eA (1989)Squnc homology btn putati ra-starch bindingdomains rom dirnt starch-dgrading nzyms. Biochm J264:309311

40. Janck S, Scik J (1999) Th olution o starch-bindingdomain. FeBS Ltt 456:119125

41. Janck S, Snsson B, MacGrgor eA (2003) Rlationbtn domain olution, spcicity, and taxonomy o th-amylas amily mmbrs containing a C-trminal starch-binding domain. eur J Biochm 270:635645

42. Rodriguz-Sanoja R, Oido N, Sanchz S (2005) Microbialstarch-binding domain. Curr Opin Microbiol 8:260267

43. Machoic M, Janck S (2006) Starch-binding domains in thpost-gnom ra. Cll Mol Li Sci 63:27102724

44. Christiansn C, Abou Hachm M, Janck S, viks-NilsnA, Blnno A, Snsson B (2009) Th carbohydrat-bindingmodul amily 20dirsity, structur, and unction. FeBS J276:50065029

45. Janck S, Snsson B, MacGrgor eA (2011) Structural and

olutionary aspcts o to amilis o non-catalytic domainsprsnt in starch and glycogn binding protins rom microbs,plants and animals. enzym Microb Tchnol 49:429440

46. Janck S (2002) Ho many consrd squnc rgionsar thr in th -amylas amily? Biologia 57(Suppl 11):2941

47. Takata H, Kuriki T, Okada S, Taksada Y, Iizuka M, MinamiuraN, Imanaka T (1992) Action o nopullulanas. Nopullulanascatalyzs both hydrolysis and transglycosylation at -(1,4)- and-(1,6)-glucosidic linkags. J Biol Chm 267:1844718452

48. Nakajima R, Imanaka T, Aiba S (1986) Comparison o aminoacid squncs o ln dirnt -amylass. Appl MicrobiolBiotchnol 23:355360

49. Toda H, Kondo K, Narita K (1982) Th complt amino acidsqunc o Taka-amylas A. Proc Jpn Acad B58:208212

50. Fridbrg F (1983) On th primary structur o amylass. FeBSLtt 152:139140

51. Rogrs JC (1985) Consrd amino acid squnc domains in-amylass rom plants, mammals, and bactria. Biochm Bio-phys Rs Commun 128:470476

52. Janck S (1992) N consrd amino acid rgion o-amylass in th third loop o thir (/)8-barrl domains. Bio-chm J 288:10691070

53. Janck S (1994) Squnc similaritis and olutionary rla-tionships o microbial, plant and animal -amylass. eur J Bio-chm 224:519524

54. MacGrgor eA, Jsprsn HM, Snsson B (1996) A circularlyprmutd -amylas-typ /-barrl structur in glucan-synth-sizing glucosyltransrass. FeBS Ltt 378:263266

55. vujicic-Zagar A, Pijning T, Kralj S, Lopz CA, euma w,Dijkhuizn L, Dijkstra Bw (2010) Crystal structur o a 117-kDa glucansucras ragmnt proids insight into olution andproduct spcicity o GH70 nzyms. Proc Natl Acad Sci USA107:2140621411

56. Ito K, Ito S, Shimamura T, wyand S, Kaarasaki Y, MisakaT, Ab K, Kobayashi T, Camron AD, Iata S (2011) Crys-tal structur o glucansucras rom th dntal caris pathognStreptococcus mutans. J Mol Biol 408:177186

57. Brison Y, Pijning T, Malbrt Y, Fabr e, Moury L, Morl S,Potocki-vrons G, Monsan P, Tranir S, Rmaud-Simon M,Dijkstra Bw (2012) Functional and structural charactrizationo -(1,2) branching sucras drid rom DSR-e glucansu-cras. J Biol Chm 287:79157924

58. Pijning T, vujicic-Zagar A, Kralj S, Dijkhuizn L, Dijkstra

Bw (2012) Structur o th -1,6/-1,4-spcic glucansucrasGTFA rom Lactobacillus reuteri 121. Acta Crystallogr Sct FStruct Biol Cryst Commun 68:14481454

59. Przylas I, Tomoo K, Trada Y, Takaha T, Fujii K, Sangr w,Strtr N (2000) Crystal structur o amylomaltas rom Ther-mus aquaticus, a glycosyltransras catalysing th productiono larg cyclic glucans. J Mol Biol 296:873886

60. Barnds TR, Bultma JB, Kapr T, an dr Maarl MJ, Dijkhui-zn L, Dijkstra Bw (2007) Thr-ay stabilization o th coa-lnt intrmdiat in amylomaltas, an -amylas-lik transgly-cosylas. J Biol Chm 282:1724217249

61. Jung JH, Jung TY, So DH, Yoon SM, Choi HC, Park BC,Park CS, woo eJ (2011) Structural and unctional analysis o


17/22


1 3

substrat rcognition by th 250s loop in amylomaltas romThermus brockianus. Protins 79:633644

62. Godany A, vidoa B, Janck S (2008) Th uniqu glycosidhydrolas amily 77 amylomaltas rom Borrelia burgdor-eri ith only catalytic triad consrd. FeMS Microbiol Ltt284:8491

63. Machoic M, Janck S (2003) Th inariant rsidus inth -amylas amily: just th catalytic triad. Biologia58:11271132

64. Jsprsn HM, MacGrgor eA, Hnrissat B, Sirks MR, Sn-sson B (1993) Starch- and glycogn-dbranching and branch-ing nzyms: prdiction o structural aturs o th catalytic(/)8-barrl domain and olutionary rlationship to othramylolytic nzyms. J Protin Chm 12:791805

65. Janck S (1997) -Amylas amily: molcular biology andolution. Prog Biophys Mol Biol 67:6797

66. Janck S, Snsson B, MacGrgor eA (2007) A rmot butsignicant squnc homology btn glycosid hydrolasclan GH-H and amily GH31. FeBS Ltt 581:12611268

67. Oslancoa A, Janck S (2002) Oligo-1,6-glucosidas andnopullulanas nzym subamilis rom th -amylas amilydnd by th th consrd squnc rgion. Cll Mol LiSci 59:19451959

68. Aghajari N, Fllr G, Grday C, Hasr R (1998) Crystal struc-turs o th psychrophilic -amylas rom Alteromonas halo-planctis in its nati orm and complxd ith an inhibitor. Pro-tin Sci 7:564572

69. Aghajari N, Fllr G, Grday C, Hasr R (1998) Structurso th psychrophilic Alteromonas haloplanctis -amylasgi insights into cold adaptation at a molcular ll.Structur 6:15031516

70. Tan TC, Mijts BN, Saminathan K, Patl BK, Din C (2008)Crystal structur o th polyxtrmophilic -amylas AmyBrom Halothermothrix orenii: dtails o a producti nzymsubstrat complx and an N domain ith a rol in binding rastarch. J Mol Biol 378:852870

71. Puspasari F, Radjasa OK, Nor AS, Nurachman Z, Syah YM,an dr Maarl M, Dijkhuizn L, Janck S, Natalia D (2013)

Ra starch-dgrading -amylas rom Bacillus aquimarisMKSC 6.2: isolation and xprssion o th gn, bioinormaticsand biochmical charactrization o th rcombinant nzym. JAppl Microbiol 114:108120

72. Bol e, Brady L, Brzozoski AM, Drnda Z, Dodson GG,Jnsn vJ, Ptrsn SB, Sit H, Thim L, woldik HF (1990)Calcium binding in -amylass: an X-ray diraction study at2.1- rsolution o to nzyms romAspergillus. Biochmis-try 29:62446249

73. vujicic-Zagar A, Dijkstra Bw (2006) Monoclinic crystal ormoAspergillus niger-amylas in complx ith maltos at 1.8 rsolution. Acta Crystallogr Sct F Struct Biol Cryst Com-mun 62:716721

74. Siddiqui KS, Poljak A, D Francisci D, Gurriro G, Pilak O,Burg D, Ratry MJ, Parkin DM, Trhlla J, Caicchioli R

(2010) A chmically modid -amylas ith a moltn-globulstat has ntropically drin nhancd thrmal stability. Protineng Ds Sl 23:769780

75. Sugahara M, Takhira M, Yutani K (2013) ect o hayatoms on th thrmal stability o -amylas rom Aspergillusoryzae. PLoS ONe 8:57432

76. Iuji H, Chino M, Kato M, Iimura Y (1996) Ra-starch-digst-ing and thrmostabl -amylas rom th yast Cryptococcussp. S-2: purication, charactrization, cloning and squncing.Biochm J 318:989996

77. Galdino AS, Ulhoa CJ, Moras LM, Prats Mv, Bloch C Jr,Torrs FA (2008) Cloning, molcular charactrization and

htrologous xprssion o AMY1, an -amylas gn romCryptococcus avus. FeMS Microbiol Ltt 280:189194

78. Styn AJ, Marmur J, Prtorius IS (1995) Cloning, squncanalysis and xprssion in yasts o a cDNA containing aLipo-myces kononenkoae-amylas-ncoding gn. Gn 166:6571

79. Kang HK, L JH, Kim D, Day DF, Robyt JF, Park KH, MoonTw (2004) Cloning and xprssion o Lipomyces starkeyi-amylas inEscherichia coli and dtrmination o som o itsproprtis. FeMS Microbiol Ltt 233:5364

80. Hostinoa e, Janck S, Gasprik J (2010) Gn squnc, bio-inormatics and nzymatic charactrization o -amylas romSaccharomycopsis fbuligera KZ. Protin J 29:355364

81. Cuyrs S, Dornz e, Dlcour JA, Courtin CM (2012) Occur-rnc and unctional signicanc o scondary carbohydratbinding sits in glycosid hydrolass. Crit R Biotchnol32:93107

82. Cockburn D, Snsson B (2013) Surac binding sits incarbohydrat acti nzyms: an mrging pictur o struc-tural and unctional dirsity. In: Lindhorst TK, Rautr AP(ds) SPR carbohydrat chmistrychmical and biologicalapproachs, ol 39. Royal Socity o Chmistry, Cambridg.doi:10.1039/9781849737173-00204

83. Mllr MS, Cockburn D, Nilsn Jw, Jnsn JM, vstr-Chris-tnsn MB, Nilsn MM, Andrsn JM, wilkns C, RannsJ, Hgglund P, Hnriksn A, Abou Hachm M, willmos M,Snsson B (2013) Surac binding sits (SBSs), mchanismand rgulation o nzyms dgrading amylopctin and -limitdxtrins. J Appl Glycosci. doi:10.5458/jag.jag.JAG-2012_023

84. Kanko A, Sudo S, Sakamoto Y, Tamura G, Ishikaa T, Ohba T(1996) Molcular-cloning and dtrmination o th nuclotid-squnc o a gn ncoding an acid-stabl -amylas romAspergillus-kawachii. J Frmnt Biong 81:292298

85. Machoic M, Janck S (2006) Th olution o putatistarch-binding domains. FeBS Ltt 580:63496356

86. Klin C, Schulz Ge (1991) Structur o cyclodxtrin glycosyl-transras rnd at 2.0 rsolution. J Mol Biol 217:737750

87. Uitdhaag JC, Mosi R, Kalk KH, an dr vn BA, DijkhuiznL, withrs SG, Dijkstra Bw (1999) X-ray structurs along thraction pathay o cyclodxtrin glycosyltransras lucidatcatalysis in th -amylas amily. Nat Struct Biol 6:432436

88. watanab K, Hata Y, Kizaki H, Katsub Y, Suzuki Y (1997) Thrnd crystal structur oBacillus cereus oligo-1,6-glucosi-das at 2.0 rsolution: structural charactrization o prolin-substitution sits or protin thrmostabilization. J Mol Biol269:142153

89. Yamamoto K, Miyak H, Kusunoki M, Osaki S (2010) Crys-tal structurs o isomaltas rom Saccharomyces cerevisiaeand in complx ith its comptiti inhibitor maltos. FeBS J277:42054214

90. Shirai T, Hung vS, Morinaka K, Kobayashi T, Ito S (2008)Crystal structur o GH13 -glucosidas GSJ rom on o thdpst sa bactria. Protins 73:126133

91. Hondoh H, Saburi w, Mori H, Okuyama M, Nakada T, Mat-

suura Y, Kimura A (2008) Substrat rcognition mchanism o-1,6-glucosidic linkag hydrolyzing nzym, dxtran glucosi-das rom Streptococcus mutans. J Mol Biol 378:913922

92. Mllr MS, Frdslund F, Majumdr A, Nakai H, Poulsn JC,Lo Lggio L, Snsson B, Abou Hachm M (2012) enzymol-ogy and structur o th GH13_31 glucan 1,6--glucosidasthat conrs isomaltooligosaccharid utilization in th probioticLactobacillus acidophilus NCFM. J Bactriol 194:42494259

93. Zhang D, Li N, Lok SM, Zhang LH, Saminathan K (2003)Isomaltulos synthas (PalI) o Klebsiella sp. LX3. Crys-tal structur and implication o mchanism. J Biol Chm278:3542835434
http://dx.doi.org/10.1039/9781849737173-00204http://dx.doi.org/10.5458/jag.jag.JAG-2012_023http://dx.doi.org/10.5458/jag.jag.JAG-2012_023http://dx.doi.org/10.1039/9781849737173-00204


18/22

. Janeek et al.

1 3

94. Raaud S, Robrt X, watzlaick H, Hasr R, Matts R,Aghajari N (2007) Trhalulos synthas nati and carbo-hydrat complxd structurs proid insights into sucrosisomrization. J Biol Chm 282:2812628136

95. Raaud S, Robrt X, watzlaick H, Hasr R, Matts R,Aghajari N (2009) Structural dtrminants o product spcic-ity o sucros isomrass. FeBS Ltt 583:19641968

96. Machius M, wigand G, Hubr R (1995) Crystal structur ocalcium-dpltdBacillus licheniormis-amylas at 2.2 rs-

olution. J Mol Biol 246:54555997. Hang KY, Song HK, Chang C, L J, L SY, Kim KK, Cho

S, St RM, Suh Sw (1997) Crystal structur o thrmostabl-amylas rom Bacillus licheniormis rnd at 1.7 rsolu-tion. Mol Clls 7:251258

98. Sud D, Fujimoto Z, Takas K, Matsumura M, Mizuno H(2001) Crystal structur o Bacillus stearothermophilus-amylas: possibl actors dtrmining th thrmostability. JBiochm 129:461468

99. Alikhajh J, Khajh K, Ranjbar B, Nadri-Mansh H, LinYH, Liu e, Guan HH, Hsih YC, Chuankhayan P, Huang YC,Jyaraman J, Liu MY, Chn CJ (2010) Structur oBacillusamyloliqueaciens -amylas at high rsolution: implicationsor thrmal stability. Acta Crystallogr Sct F Struct Biol CrystCommun 66:121129

100. Fujimoto Z, Takas K, Doui N, Momma M, Matsumoto T,Mizuno H (1998) Crystal structur o a catalytic-sit mutant-amylas rom Bacillus subtilis complxd ith maltopnta-os. J Mol Biol 277:393407

101. Dais GJ, Brzozoski AM, Dautr Z, Rasmussn MD,Borchrt Tv, wilson KS (2005) Structur o a Bacillus halma-palus amily 13 -amylas, BHA, in complx ith an acarbos-drid nonasaccharid at 2.1 rsolution. Acta Crystallogr DBiol Crystallogr 61:190193

102. Lyhn-Irsn L, Hobly TJ, Kaasgaard SG, Harris P (2006)Structur oBacillus halmapalus -amylas crystallizd ithand ithout th substrat analogu acarbos and maltos. ActaCrystallogr Sct F Struct Biol Cryst Commun 62:849854

103. Nonaka T, Fujihashi M, Kita A, Hagihara H, Ozaki K, Ito S,

Miki K (2003) Crystal structur o calcium-r -amylasrom Bacillus sp. strain KSM-K38 (AmyK38) and its sodiumion binding sits. J Biol Chm 278:2481824824

104. Shirai T, Igarashi K, Ozaa T, Hagihara H, Kobayashi T, OzakiK, Ito S (2007) Ancstral squnc olutionary trac and crys-tal structur analyss o alkalin -amylas rom Bacillus sp.KSM-1378 to clariy th alkalin adaptation procss o pro-tins. Protins 66:600610

105. Kanai R, Haga K, Akiba T, Yaman K, Harata K (2004) Bio-chmical and crystallographic analyss o maltohxaos-pro-ducing amylas rom alkalophilicBacillus sp. 707. Biochmis-try 43:1404714056

106. Tsukamoto A, Kimura K, Ishii Y, Takano T, Yaman K (1988)Nuclotid squnc o th maltohxaos-producing amyl-as gn rom an alkalophilic Bacillus sp. #707 and structural

similarity to liquying typ -amylass. Biochm Biophys RsCommun 151:2531107. an dr Kaaij RM, Janck S, an dr Maarl MJ, Dijkhuizn

L (2007) Phylogntic and biochmical charactrization o anol clustr o intracllular ungal -amylas nzyms. Micro-biology 153:40034015

108. Marion CL, Rapply CA, engl JT, Goldman we (2006)An -(1,4)-amylas is ssntial or -(1,3)-glucan produc-tion and irulnc in Histoplasma capsulatum. Mol Microbiol62:970983

109. Camacho e, Spulda ve, Goldman we, San-Blas G, Nino-vga GA (2012) exprssion o Paracoccidioides brasiliensisAMY1 in a Histoplasma capsulatum amy1 mutant, rlats an

-(1,4)-amylas to cll all -(1,3)-glucan synthsis. PLoSONe 7:50201

110. Rodriguz Sanoja R, Morlon-Guyot J, Jor J, Pintado J, Jug N,Guyot JP (2000) Comparati charactrization o complt andtruncatd orms oLactobacillus amylovorus -amylas androl o th C-trminal dirct rpats in ra-starch binding. Appleniron Microbiol 66:33503356

111. Rodriguz-Sanoja R, Ruiz B, Guyot JP, Sanchz S (2005)Starch-binding domain acts catalysis in to Lactobacillus-amylass. Appl eniron Microbiol 71:297302

112. Guilln D, Santiago M, Linars L, Prz R, Morlon J, Ruiz B,Sanchz S, Rodriguz-Sanoja R (2007) -Amylas starch bind-ing domains: cooprati cts o binding to starch granulso multipl tandmly arrangd domains. Appl eniron Micro-biol 73:38333837

113. Rodriguz-Sanoja R, Oido N, escalant L, Ruiz B, SanchzS (2009) A singl rsidu mutation abolishs attachmnt o thCBM26 starch-binding domain romLactobacillus amylovorus-amylas. J Ind Microbiol Biotchnol 36:341346

114. Janck S, Lqu e, Blarbi A, Hay B (1999) Clos olu-tionary rlatdnss o -amylass rom Archaa and plants. JMol eol 48:421426

115. Da Lag JL, Fllr G, Janck S (2004) Horizontal gntransr rom eukarya to bactria and domain shufing: th-amylas modl. Cll Mol Li Sci 61:97109

116. Jons RA, Jrmiin LS, eastal S, Patl BK, Bacham IR (1999)Amylas and 16S rRNA gns rom a hyprthrmophilicarchabactrium. J Appl Microbiol 86:93107

117. Lqu e, Janck S, Blarbi A, Hay B (2000) Thrmophilicarchaal amylolytic nzyms. enzym Microb Tchnol 26:213

118. Lqu e, Hay B, Blarbi A (2000) Cloning and xprssiono an -amylas ncoding gn rom th hyprthrmophilicarchabactrium Thermococcus hydrothermalis and biochmi-cal charactrisation o th rcombinant nzym. FeMS Micro-biol Ltt 186:6771

119. Lim JK, L HS, Kim YJ, Ba SS, Jon JH, Kang SG, L JH(2007) Critical actors to high thrmostability o an -amylasrom hyprthrmophilic archaon Thermococcus onnurineusNA1. J Microbiol Biotchnol 17:12421248

120. Dong G, viill C, Sachnko A, Zikus JG (1997) Cloning,squncing, and xprssion o th gn ncoding xtracllular-amylas rom Pyrococcus uriosus and biochmical charac-trization o th rcombinant nzym. Appl eniron Microbiol63:35693576

121. Jorgnsn S, vorgias Ce, Antranikian G (1997) Cloning,squncing, charactrization, and xprssion o an xtracllular-amylas rom th hyprthrmophilic archaon Pyrococcusuriosus in Escherichia coli andBacillus subtilis. J Biol Chm272:1633516342

122. Frillingos S, Lindn A, Nihaus F, vargas C, Nito JJ, vntosaA, Antranikian G, Drainas C (2000) Cloning and xprssion o-amylas rom th hyprthrmophilic archaon Pyrococcuswoesei in th modratly halophilic bactriumHalomonas elon-

gata. J Appl Microbiol 88:495503123. Lindn A, Mayans O, Myr-Klauck w, Antranikian G, wil-manns M (2003) Dirntial rgulation o a hyprthrmophilic-amylas ith a nol (Ca,Zn) to-mtal cntr by zinc. JBiol Chm 278:98759884

124. Lindn A, wilmanns M (2004) Adaptation o class-13-amylass to dirs liing conditions. ChmBioChm5:231239

125. Rogrs JC, Milliman C (1983) Isolation and squncanalysis o a barly -amylas cDNA clon. J Biol Chm258:81698174

126. Rogrs JC (1985) To barly -amylas gn amilis ar rgu-latd dirntly in aluron clls. J Biol Chm 260:37313738


19/22


1 3

127. Kadziola A, Ab J, Snsson B, Hasr R (1994) Crys-tal and molcular structur o barly -amylas. J Mol Biol239:104121

128. Kadziola A, Sgaard M, Snsson B, Hasr R (1998) Molcularstructur o a barly -amylasinhibitor complx: implicationsor starch binding and catalysis. J Mol Biol 278:205217

129. Robrt X, Hasr R, Gottschalk Te, Ratajczak F, DriguzH, Snsson B, Aghajari N (2003) Th structur o bar-ly -amylas isozym 1 rals a nol rol o domain C

in substrat rcognition and binding: a pair o sugar tongs.Structur 11:973984

130. Robrt X, Hasr R, Mori H, Snsson B, Aghajari N (2005)Oligosaccharid binding to barly -amylas 1. J Biol Chm280:3296832978

131. Baulcomb DC, Huttly AK, Martinssn RA, Barkr RF, JarisMG (1987) A nol hat -amylas gn (-Amy3). Mol GnGnt 209:3340

132. Huang N, Sutli TD, Litts JC, Rodriguz RL (1990) Classica-tion and charactrization o th ric -amylas multign am-ily. Plant Mol Biol 14:655668

133. Young Te, DMason DA, Clos TJ (1994) Cloning o an-amylas cDNA rom aluron tissu o grminating maizsd. Plant Physiol 105:759760

134. Mori H, Kobayashi T, Tonokaa T, Tatmatsu A, Matsui H,Kimura A, Chiba S (1997) Molcular cloning o an -amylascDNA rom grminating cotyldons o kidny ban (Phaseolusvulgaris L. c. Toramam). J Appl Glycosci 45:261267

135. wgrzyn T, Rilly K, Cipriani G, Murphy P, Ncomb R, Gard-nr R, MacRa e (2000) A nol -amylas gn is transintlyuprgulatd during lo tmpratur xposur in appl ruit. eurJ Biochm 267:13131322

136. Stanly D, Fitzgrald AM, Farndn KJA, MacRa eA (2002)Charactrisation o putati -amylass rom appl (Malusdomestica) and Arabidopsis thaliana. Biologia 57(Suppl11):137148

137. Junior Av, do Nascimnto JR, Lajolo FM (2006) Molcularcloning and charactrization o an -amylas occurring in thpulp o ripning bananas and its xprssion in Pichia pastoris. J

Agric Food Chm 54:82228228138. Bozonnt S, Jnsn MT, Nilsn MM, Aghajari N, JnsnMH, Kramht B, willmos M, Tranir S, Hasr R, SnssonB (2007) Th pair o sugar tongs sit on th non-catalyticdomain C o barly -amylas participats in substrat bindingand actiity. FeBS J 274:50555067

139. Nilsn MM, Bozonnt S, So eS, Motyan JA, Andrsn JM,Dilokpimol A, Abou Hachm M, Gymant G, Nastd H, Kan-dra L, Sigurskjold Bw, Snsson B (2009) To scondarycarbohydrat binding sits on th surac o barly -amylas1 ha distinct unctions and display synrgy in hydrolysis ostarch granuls. Biochmistry 48:76867697

140. Nilsn Jw, Kramht B, Bozonnt S, Abou Hachm M, StippSL, Snsson B, willmos M (2012) Dgradation o th starchcomponnts amylopctin and amylos by barly -amylas 1:

rol o surac binding sit 2. Arch Biochm Biophys 528:16141. Tranir S, Dill K, Robrt X, Bozonnt S, Hasr R, SnssonB, Aghajari N (2005) Insights into th pair o sugar tongssurac binding sit in barly -amylas isozyms and crystal-lization o appropriat sugar tongs mutants. Biologia 60(Suppl16):3746

142. Pujadas G, Palau J (2001) eolution o-amylass: architc-tural aturs and ky rsidus in th stabilization o th (/)8scaold. Mol Biol eol 18:3854

143. Godany A, Majzloa K, Horathoa v, vidoa B, JanckS (2010) Tyrosin 39 o GH13 -amylas rom Thermococ-cus hydrothermalis contributs to its thrmostability. Biologia65:408415

144. Da Lag JL, Danchin eG, Casan D (2007) whr do ani-mal -amylass com rom? An intrkingdom trip. FeBS Ltt581:39273935

145. Fllr G, Lonhinn T, Droann C, Libioull C, van BumnJ, Grday C (1992) Purication, charactrization, and nuclo-tid squnc o th thrmolabil -amylas rom th Antarc-tic psychrotroph Alteromonas haloplanctis A23. J Biol Chm267:52175221

146. Fllr G, Payan F, Thys F, Qian M, Hasr R, Grday C (1994)

Stability and structural analysis o-amylas rom th Antarc-tic psychrophilAlteromonas haloplanctis A23. eur J Biochm222:441447

147. DAmico S, Grday C, Fllr G (2000) Structural similari-tis and olutionary rlationships in chlorid-dpndnt-amylass. Gn 253:95105

148. Aghajari N, Fllr G, Grday C, Hasr R (2002) Structural basiso-amylas actiation by chlorid. Protin Sci 11:14351441

149. Cipolla A, Dlbrassin F, Da Lag JL, Fllr G (2012) Tmpr-atur adaptations in psychrophilic, msophilic and thrmophilicchlorid-dpndnt -amylass. Biochimi 94:19431950

150. Bor PH, Hicky DA (1986) Th -amylas gn inDrosophilamelanogaster: nuclotid squnc, gn structur and xprs-sion motis. Nuclic Acids Rs 14:83998411

151. Da Lag JL, wgnz M, Cariou ML (1996) Distribution andolution o introns in Drosophila amylas gns. J Mol eol43:334347

152. Da Lag JL, Rnard e, Chartois F, Lmunir F, Cariou ML(1998) Amyrl, a paralogous gn o th amylas gn am-ily in Drosophila melanogasterand th Sophophora subgnus.Proc Natl Acad Sci USA 95:68486853

153. Da Lag JL, Maisonhaut C, Maczkoiak F, Cariou ML (2003)A nstd -amylas gn inDrosophila ananassae. J Mol eol57:355362

154. Strobl S, Gomis-Ruth FX, Maskos K, Frank G, Hubr R,Glockshubr R (1997) Th -amylas rom th yllo malorm: complt primary structur, crystallization and prlimi-nary X-ray analysis. FeBS Ltt 409:109114

155. Strobl S, Maskos K, Btz M, wigand G, Hubr R, Gomis-Ruth

FX, Glockshubr R (1998) Crystal structur o yllo malorm -amylas at 1.64 rsolution. J Mol Biol 278:617628156. L Moin S, Sllos D, Moal J, Danil JY, San Juan Srrano F,

Samain JF, van wormhoudt A (1997) Amylas on Pecten maxi-mus (Mollusca, bials): protin and cDNA charactrization;quantication o th xprssion in th digsti gland. Mol MarBiol Biotchnol 6:228237

157. Nikapitiya C, Oh C, whang I, Kim CG, L YH, Kim SJ, LJ (2009) Molcular charactrization, gn xprssion analysisand biochmical proprtis o-amylas rom th disk abalon,Haliotis discus discus. Comp Biochm Physiol B Biochm MolBiol 152:271281

158. van wormhoudt A, Sllos D (1996) Cloning and squncinganalysis o thr amylas cDNAs in th shrimp Penaeus van-namei (Crustaca dcapoda): olutionary aspcts. J Mol eol

42:543551159. Da Lag JL, Maczkoiak F, Cariou ML (2011) Phylognticdistribution o intron positions in -amylas gns o bilatriasuggsts numrous gains and losss. PLoS ONe 6:19673

160. Buisson G, Du e, Hasr R, Payan F (1987) Thr-dimn-sional structur o porcin pancratic -amylas at 2.9 rsolution. Rol o calcium in structur and actiity. eMBO J6:39093916

161. Qian M, Hasr R, Payan F (1993) Structur and molcularmodl rnmnt o pig pancratic -amylas at 2.1 rsolu-tion. J Mol Biol 231:785799

162. Qian M, Hasr R, Buisson G, Du e, Payan F (1994) Thacti cntr o a mammalian -amylas. Structur o th


20/22

. Janeek et al.

1 3

complx o a pancratic -amylas ith a carbohydrat inhibi-tor rnd to 2.2- rsolution. Biochmistry 33:62846294

163. Gills C, Astir JP, Marchis-Mourn G, Cambillau C, Payan F(1996) Crystal structur o pig pancratic -amylas isonzymII, in complx ith th carbohydrat inhibitor acarbos. eur JBiochm 238:561569

164. Ramasubbu N, Paloth v, Luo Y, Brayr GD, Lin MJ (1996)Structur o human saliary -amylas at 1.6 Angstrom rsolu-tion: implications or its rol in th oral caity. Acta Crystallogr

D 52:435446165. Brayr GD, Luo Y, withrs SG (1995) Th structur o human

pancratic -amylas at 1.8 rsolution and comparisons ithrlatd nzyms. Protin Sci 4:17301742

166. Mizutani K, Toyoda M, Otak Y, Yoshioka S, Takahashi N,Mikami B (2012) Structural and unctional charactrization orcombinant mdaka sh -amylas xprssd in yast Pichiapastoris. Biochim Biophys Acta 1824:954962

167. Machius M, vrtsy L, Hubr R, wigand G (1996) Carbo-hydrat and protin-basd inhibitors o porcin pancratic-amylas: structur analysis and comparison o thir bindingcharactristics. J Mol Biol 260:409421

168. Payan F, Qian M (2003) Crystal structur o th pig pancratic-amylas complxd ith malto-oligosaccharids. J ProtinChm 22:275284

169. Larson SB, Day JS, McPhrson A (2010) X-ray crystallographicanalyss o pig pancratic -amylas ith limit dxtrin, oligo-saccharid, and -cyclodxtrin. Biochmistry 49:31013115

170. Qin X, Rn L, Yang X, Bai F, wang L, Gng P, Bai G, ShnY (2011) Structurs o human pancratic -amylas in complxith acariostatins: implications or drug dsign against typ IIdiabts. J Struct Biol 174:196202

171. Yang CH, Liu wH (2007) Cloning and charactrization oa maltotrios-producing -amylas gn rom Thermobifdausca. J Ind Microbiol Biotchnol 34:325330

172. Doukyu N, Yamagishi w, Kuahara H, Ogino H (2008) Amaltooligosaccharid-orming amylas gn rom Brachybac-terium sp. strain LB25: cloning and xprssion in Escherichiacoli. Biosci Biotchnol Biochm 72:24442447

173. Yamaguchi R, T

2013 doi ?-amylase an enzyme specificity found in various families of glycoside hydrolases

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