Parnas pathway from archaea and hyperthermophilic bacteria suppo

24 Pages·2014·1.78 MB·English

Archaea1, 199–221
© 2003 Heron Publishing—Victoria, Canada
Distribution and phylogenies of enzymes of the Embden-Meyerhof-
Parnas pathway from archaea and hyperthermophilic bacteria support
a gluconeogenic origin of metabolism
1,2 1
RON S. RONIMUS and HUGH W. MORGAN
1 ThermophileResearch Unit, Department of Biological Sciences, University ofWaikato, Private Bag 3105, Hamilton, New Zealand
2 Author to whom correspondence should be addressed ([email protected])
Received June 17, 2002; accepted October 24, 2002; published online November 28, 2002
Summary Enzymes of the gluconeogenic/glycolytic path- the Oparin-Haldane model or prebiotic souphypothesis(ex-
way(theEmbden-Meyerhof-Parnas(EMP)pathway),there- poundedbyStanleyMillerandcolleagues);(2)thepyruvate-
ductive tricarboxylic acid cycle, the reductive pentose phos- pulledchemoautotrophicoriginofmetabolismonthesurface
phate cycle and the Entner-Doudoroff pathway are widely ofmetalsulfideminerals(Wächtershäuser1988a,1988b);and
distributedandareoftenconsideredtobecentraltotheorigins (3) thetheoryofextraterrestrialseedingofEarth,duringthe
ofmetabolism.Inparticular,severalenzymesofthelowerpor- earlyformativeyears,withorganiccompounds(reviewedby
tionoftheEMPpathway(theso-calledtrunkpathway),includ- Orgel1998andMaden1995).TheWächtershäusertheoryen-
ingtriosephosphateisomerase(TPI;EC5.3.1.1),glyceralde- visages a hyperthermophilic and autotrophic origin of life,
hyde-3-phosphatedehydrogenase(GAPDH;EC1.2.1.12/13), whereastheOparin-Haldaneandextraterrestrialseedingtheo-
phosphoglyceratekinase(PGK;EC2.7.2.3)andenolase(EC ries presuppose a heterotrophic lifestyle (Wächtershäuser
4.2.1.11), are extremely well conserved and universally dis- 1988a, 1988b, 1994). In addition, the notion of an “RNA
tributedamongthethreedomainsoflife.Inthispaper,thedis- world,” in which RNA-based life preceded DNA-based life,
tributionofenzymesofgluconeogenesis/glycolysisinhyper- hasgainedwideacceptance,althoughithasdetractors.How-
thermophiles—microorganisms that many believe represent ever,evenaputativeRNA-based“organism”couldhavearisen
theleastevolvedorganismsontheplanet—isreviewed.Inad- onlyfromaprebioticchemicalenvironmentconducivetoits
dition,thephylogeniesofthetrunkpathwayenzymes(TPIs, existence (Poole et al. 1999).
GAPDHs, PGKs and enolases) are examined. The enzymes Although these theories and their associated predictions
catalyzingeachofthesix-carbontransformationsintheupper havedonemuchtoprovideexplanationsforthepotentialarray
portion of the EMP pathway, with the possible exception of ofprebioticchemistries,itisuncertainwhichoneofthethree
aldolase,areallderivedfrommultiplegenesequencefamilies. bestexplainstheoriginoflife.However,therearecluesthat
In contrast, single sequence families can account for the canbegleanedfromthedistributionofkeyenzymesofmetab-
archaealandhyperthermophilicbacterialenzymeactivitiesof olisminpresent-dayorganismsandfromphylogeneticanaly-
thelowerportionoftheEMPpathway.Theuniversaldistribu- sisofuniversallydistributedenzymes.Asimilarapproachhas
tionofthetrunkpathwayenzymes,incombinationwiththeir beenusedwithDNA-dependentRNApolymerases(Klenket
phylogenies, supports the notion that the EMP pathway al. 1994), translation factors EF-Tu and EF-G, ATPase sub-
evolvedinthedirectionofgluconeogenesis,i.e.,fromthebot- units, tRNA (tRNA-Met-E and tRNA Met-I; Iwabe et al.
tom up. 1989), aminoacyl-tRNA synthetases (Brown and Doolittle
1995),RNApolymerases(Iwabeetal.1991)andheat-shock
Keywords: enolase, evolution, gluconeogenesis, glyceralde-
proteins (Gupta 1998).
hyde-3-phosphate dehydrogenase, glycolysis, origins of life,
Universalphylogeniesbasedonsmallsubunitribosomalse-
phosphoglyceratekinase,triosephosphateisomerase.
quence comparisons have been cited as evidence that ex-
tremely thermophilic (optimal growth temperatures above
70 °C) and hyperthermophilic microorganisms (optimal
Introduction
growth temperatures above 80 °C) are the least evolved of
Modern day reconstruction of the ancient chemical and bio- present-day microorganisms, and therefore most closely re-
logicalscenariosthatledtotheoriginsoflifeonEarthapprox- semble the last common universal ancestor (Woese et al.
imately3.8billionyearsagoisaninherentlydifficulttaskand 1990).Thus,hyperthermophilesrepresentthemostappropri-
isintimatelytiedtotheoriginsofmetabolism.Therearethree ate microorganisms with which to investigate facets of the
generaltheoriesorhypothesesthatattempttodelineatethefa- earlyevolutionoflifeandmetabolism.Inaccordancewiththe
vorableconditionsinwhichlifeispresumedtohavearisen:(1) “thermophiles-first” hypothesis, deep sea hydrothermal vent
200 RONIMUS AND MORGAN
ecosystemshavebeensuggestedtorepresentthemostancient, the PEDANT database (http://pedant.gsf.de), the COG data-
continuously inhabited microbial habitats on Earth (Reysen- base (Tatusov et al. 2001), the BRENDA enzyme database
bachandShock2002).Inaddition,geologicalevidencesup- (BRENDA:http://www.brenda.uni-koeln.de),conserveddo-
portstheviewthathotaquatichabitatsexistedthroughoutthe main architecture retrieval toll (CDART at NCBI) and the
early history of Earth, regardless of the temperature of the KEGGenzymaticpathwaydatabase(http://www.genome.ad.
ocean(circa4.3Gayearsago)oroftheatmosphere(Baross jp/kegg/metabolism.html). Sequence alignments and phylo-
1998).Itshouldbenoted,however,thatthenotionthatthefirst genetictreeswereproducedwithfull-lengthsequencesasre-
microorganisms were hyperthermophiles is not universally portedinthedatabasesusingtheneighbor-joiningmethodof
accepted.Forexample,othershavearguedthathyperthermo- ClustalW 1.7 and ClustalX 1.81, unless otherwise stated
philesmayhaveevolvedfrommesophilicancestors,basedon (SaitouandNei1987,Thompsonetal.1994;forbrevity,the
comparisonoftheenzymereversegyrase,whichisfoundinall alignmentsarenotshown).Thealignmentswereexaminedfor
organisms with optimum temperatures over 70 °C (Forterre accuracy by comparison with secondary structure in the se-
1996).Furthermore,ahyperthermophilicoriginoflifeisalso quences derived from crystal structures in the Protein Data
contrarytotheRNAworldhypothesisbecauseoftherelative Base (PDB).
instability of RNA at high temperatures (Poole et al. 1999). For maximum parsimony analyses, the alignments were
Toacquireaclearerunderstandingoftheoriginsofgluco- firstformattedforPAUP*(PhylogeneticAnalysisUsingParsi-
neogenesis/glycolysis,itisinformativetoexaminethedistri- mony(*andothermethods))usingtheWebPhylipClustalcon-
bution and phylogenetic origins of the enzymes of gluco- verter(LimandZhang1999).Maximumparsimonyanalyses
neogenesis/glycolysis and their associated biochemical wereconductedwithPAUP*4.0Beta10Versionbyperform-
characteristics. This, in turn, could help provide supporting ingheuristicsearcheswithrandomstepwise-addition(10ad-
evidenceforoneormoreofthecompetingevolutionarytheo- dition-sequence replicates) with the branch swapping algo-
riesofmetabolicdevelopmentmentionedabove.Thecentral rithm tree-bisection-reconnection (Swofford et al. 1996,
metabolicpathwayofglycolysisispresentineachofthethree Swofford 2002). When more than one maximum parsimony
domains, although there is significant variation in enzymes, treewassaved,usingthestrictguidelinesoption,aconsensus
especially in the Archaea and in hyperthermophilic bacteria tree was generated and statistically validated with 500 boot-
(Fothergill-GilmoreandMichels1993,SchönheitandSchäfer straps. Statistical validation of the tree branch nodes and
1995, Selig et al. 1997, Ronimus and Morgan 2001). lengthsforallphylogenieswereassessedusing500bootstraps
Several reviews of the distribution of glycolytic enzymes and values were rounded to % values. The triosephosphate
werepublishedin1999orearlier,butwiththerapidincreasein isomerase(TPI),glyceraldehyde-3-phosphatedehydrogenase
genome sequencing, resulting in 10 new published archaeal (GAPDH),phosphoglyceratekinase(PGK)andenolaseanal-
genomes(Fothergill-GilmoreandMichels1993,Dansonetal. ysesutilized282,466,415and464phylogeneticallyinforma-
1998,Galperinetal.1998a,Cordwell1999,Dandekaretal. tive amino acid residues, respectively. The number of se-
1999,Koikeetal.1999),sufficientprogresshasbeenmadeto quences available in the database was estimated using the
warrant the new summary presented in this paper. Further- BLASTP (non-redundant database with daily updates) and
more,themostrecentoftheseearlierreviews(1998–1999)ex- Pfam databases (Altschul et al. 1997, Sonnhammer et al.
aminedonlythedistributionofenzymes,notthephylogenetic 1997). Quoted % similarities and % identities were deter-
relationshipsbetweenenzymesofrepresentativeorganismsof minedusingpraze(http://www.tigr.org)withalignmentbythe
the three domains. In this paper, we summarize the current Smith-Watermanalgorithm,oralternativelywiththeBLAST
state of knowledge regarding (1) enzymes of glycolysis/ two sequence option (Altschul et al. 1997).
gluconeogenesis among hyperthermophilic microorganisms,
both bacterial and archaeal, and (2) fully sequenced meso-
philicandthermophilicarchaealgenomes.Inaddition,weex-
Results
aminetheevolutionoftheuniversallydistributedenzymesof
the lower trunk pathway of glycolysis. Our results provide
strong support for a gluconeogenic origin for the Embden- Distribution of glycolytic/gluconeogenic enzymes in
Meyerhof-Parnas (EMP) pathway. hyperthermophiles
Thedistributionofgenesencodingglycolytic/gluconeogenic
Materials and methods enzymes, their Enzyme Commission (EC) numbers and
four-digitidentificationnumbersordatabaseaccessioncodes
Data analysis and phylogenetic tree construction
are shown in Table 1. Individual enzymes and the gene se-
Representativeaminoacidsequencesfromtheglycolyticen- quence families that encode them will be discussed sequen-
zyme gene families were used in BLASTP, Gapped-BLAST tiallystartingwiththefirststepofglycolysis,whichiscata-
andPSI-BLASTsearchesofthenon-redundantdatabase(Alt- lyzed by hexokinases/glucokinases. Table 2 summarizes the
schuletal.1990,1997)andretrievedforanalysis.Inaddition, enzymatic activities associated with cell-free extracts of the
weusedtheComprehensiveMicrobialResourcewiththegene organisms presented in Table 1 or with purified or heterolo-
matrix option at the Institute for Genome Research (TIGR: gouslyexpressedproteinsfromtypestrainsorfromcloselyre-
http://www.tigr.org/tdb/mdb/mdbcomplete.html), as well as lated species or strains.
ARCHAEA VOLUME 1, 2003
EMBDEN-MEYERHOF-PARNAS PATHWAY IN HYPERTHERMOPHILES 201
Glucokinases naschii(MJ1605;Cordwell1999).WhentheAeropyrumper-
nixgeneAPE0768(relatedtotheannotatedAquifexaeolicus
Bacterial glucokinases (GLKs; EC 2.7.1.2; ATP:D-glucose-
PGI gene AA0750; Cordwell 1999) was used in a BLASTP
6-phosphotransferase)preferentiallycatalyzethephosphoryl-
sequencesearch,orthologstotheclassicannotatedPGIwere
ationofglucoseintheEMPpathway,withhighspecificity.Se-
foundinthecrenarchaeotesPyrobaculumaerophilum(strain
quence comparisons have shown that bacterial glucokinases
IM2; PAE1610), S. solfataricus (SSO2281), S. tokodaii
have no apparent homology to the less specific hexokinases
(ST2245), T. acidophilum (TA1419) and T. volcanium
foundthroughoutthehigherEukarya(EC2.7.1.1;Cárdenaset
(TV1438;Altschuletal.1997).Arelatedsequencewasalso
al.1998).However,apotentialancestralrelationshiphasbeen
foundinM.thermoautotrophicum(MTH1546usingtheCOG
suggested(Cárdenasetal.1998)onthebasisofthesizeofthe
database; Tatusov et al. 2001), but not in other archaeal
bacterial enzyme, which is about half that of the larger eu-
genomes.
karyal protein.
The gene encoding the PGI from Pyrococcus furiosus
Glucokinasesequencesfallintofourrelatedsubgroups:glk,
(DSM3638)hasrecentlybeenclonedandsequenced(Hansen
yajF,yhc1andyeast-typeGLKs.TheAquifexaeolicusstrain
andSchönheit2001b,Verheesetal.2001b).TheP.furiosusse-
VF5enzymeisoftheyajFtype(Dandekaretal.1999).Aero-
quencepossessesonly3–6%identitywiththeclassicPGIen-
pyrum pernix strain K1, T. acidophilum (DSM 1728), Halo-
zyme family, and evidence from structure-based sequence
bacterium strain NRC-1 and T. volcanium strain GSS1 each
analysisindicatesthatthesePGIsareunlikelytobephylogen-
possessan annotated GLK of the yhc1 sequence type (Dan-
eticallyrelated(HansenandSchönheit2001b,Verheesetal.
dekaretal.1999),whichshare37,33,32and35%sequence
2001b).WhentheP.furiosussequence(PF0196)wasusedto
identitywiththeHaemophilusinfluenzaeGLKsequence,re-
probe the database, similar sequences were found in the
spectively(TIGR:http://www.tigr.org/tdb/mdb/mdbcomplete.
P. horikoshii (strain OT3; PH1956), P. abysii (strain GE5;
html).
PA1199)andunfinishedMethanosarcinabarkeriiandMeth-
Adenosine diphosphate-dependent GLKs (EC 2.7.1.147)
anosarcinamazeigenomes,butnotinthegenomeofArchaeo-
are found in Pyrococcus and Thermococcus species, and in
globus fulgidus strain VC16. Lower scoring, paralogous
Methanococcus (Methanocaldococcus; DSM 2661) jan-
sequences are present in P. furiosus, P. abysii, P. horikoshii,
naschii(Kengenetal.1994,1995,Seligetal.1997,Kogaetal.
Methanosarcina acetivorans(strain C2A) and M. jannaschii
2000, Ito et al. 2001, Sakuraba et al. 2002, Verhees et al.
(Table 1; Altschul et al. 1997, Verhees et al. 2001b). Using
2002b).TheseGLKsareunrelatedtotheATP-dependentglu-
PSI-BLAST with a second iteration cycle, an ortholog to
cokinasesandhexokinases.However,structuralcomparisons
PF0196 was found in Methanopyrus kandleri that probably
indicatethattheyaredistantlyrelatedtoFamilyBsugarkin-
encodesabonafidePGI(strainAV19;Table1).Thus,thecur-
ases similar to ribokinase (Ito et al. 2001). The monomeric
rentevidenceindicatesthepresenceofatleasttwosequence
protein from M. jannaschii is bifunctional, containing both
familiesforPGIs(forspeciesorstrainsinwhichPGIactivity
ADP-dependentglucokinaseandphosphofructokinaseactivi-
has been found; see Table 2).
ties,withtheglucokinasereactionratebeingthreefoldhigher
than the phosphofructokinase reaction rate (Sakuraba et al.
Phosphofructokinases
2002). No gene sequence has been identified for a GLK in
S.solfataricus(strainP2),althoughGLKactivityhasbeende- Phosphofructokinases (PFKs) catalyze the third step of gly-
scribed (De Rosa et al. 1984). No GLK has been identified colysis:phosphorylationoffructose-6-phosphateatthe1posi-
through sequence analysis in Archaeoglobus fulgidus strain tion(recentlyreviewedbyRonimusandMorgan2001).The
VC16 (DSM 4304), although ADP-glucokinase activity has well-characterized,textbook,FamilyAATP-dependentPFKs
been demonstrated in A. fulgidus strain 7324 (Labes and (ATP-PFKs;EC2.7.1.11),whichcatalyzeanirreversiblereac-
Schönheit 2001). tion,andthepyrophosphate-dependentPFKs(PPi-PFKs;EC
Thus,thecurrentevidencedemonstratesthepresenceofat 2.7.1.90),whichcatalyzeareversiblereaction,arewidelydis-
leasttwounrelatedsequencefamiliesforglucokinases/hexo- tributed among bacteria and eukaryotes. The PPi-PFKs are
kinases,andmostlikelythree,basedonthelackofaconvinc- found predominantly in anaerobic bacteria, some plants and
ing common ancestral sequence for bacterial GLKs and the lowereukaryotes,butalsointhecrenarchaeoteThermoproteus
hexokinases found in most higher eukaryotes. tenax (Morgan and Ronimus 1998, Siebers et al. 1998,
RonimusandMorgan2001).ThermotogahasbothaFamilyA
ATP-PFK, allostericallyregulatedbyPP andpolyphosphate
i
Phosphoglucoisomerase
(Dingetal.2001),andaPP-PFK.TheATP-PFKisalsoregu-
i
Phosphoglucoisomerase (PGI; EC 5.3.1.9), also called glu- lated by typical bacterial effectors such as ADP (activation)
cose-6-phosphateisomerase,catalyzesthesecondstepinthe andphosphoenolpyruvate(inhibition;Hansenetal.2002).Se-
glycolyticpathway:thereversiblealdose/ketoseisomerization quence analysis also indicates that a Family A ATP-PFK is
of glucose-6-phosphate to fructose-6-phosphate (Fothergill- presentinAquifexaeolicus(AE1708).ArecentiterativePSI-
GilmoreandMichels1993).AclassicPGIgene(homologous BLASTsequenceanalysissuggeststhatdiacylglyceridekin-
totheE.coliPGI)hasbeenidentifiedinthehyperthermophilic ases,sphingosinekinasesandNADkinasesmaybedistantly
bacteriumThermotogamaritima(strainMSB8;TM1385)and related to PFKs and may share a similar phosphate-donor
alsointheeuryarchaeotesHalobacteriumNRC-1andM.jan- binding site and reaction mechanism (Labesse et al. 2002).
ARCHAEA ONLINE at http://archaea.ws
202 RONIMUS AND MORGAN
archaeathatencodeglycolyticenzymes.GAP=glyceraldehyde-3-phosphatede-hosphoglyceratemutase;PK=pyruvate EnolasePKPPDK,PEPsyn 4.2.1.112.7.1.402.7.9.1,2.7.9.2 0484–2142 087702080272 2458048906500026 081208192423 1132–0710 112614410057 021511880043 194205700092 gi10580680gi10579952PpsA 023201080542 1647–0252 16723890340826672458 .Continuedonfacingpage
ce;p
mophilesandmesophiliosephosphateisomerashoglycerate-dependent iPGM/dPGM 35.4.2.1 17441990d0542 1374d1774 d1616 d2326 d1751d1425 d2318 d1959 d0037 d80748gi10581330 d1612d0010 d1193 d01323998
ypertherTPI=tribisphosp PGK 2.7.2. 0118 0689 0173 1742 1146 1679j 1057j 1218 gi105 0641 1662 35922669
mbersofgenesidentifiedinhfructose-1,6-bisphosphatase;ceratemutase;dPGM=2,3- TPIGAP 5.3.1.11.2.1.12 03601065 06890688 15380171 15011740 13041732 12080257 19201874 18841830 gi10580581gi10579745 15281146 16640618 460710183345
Table1.EnzymeCommission(EC)numbersandfour-digitidentificationnumbersordatabaseaccessionnuAbbreviations:GLK=glucokinase;PGI=phosphoglucoisomerase;PFK=phosphofructokinase;FBPase=hydrogenase;PGK=phosphoglyceratekinase;iPGM=2,3-bisphosphoglycerate-independentphosphoglykinase;PPDK=pyruvate:orthophosphatedikinase;andPEPsyn=phosphoenolpyruvatesynthase. GLKPGIPFKFBPaseAldolase ECnumber(s)2.7.1.15.3.1.92.7.1.113.1.3.114.1.2.132.7.1.22.7.1.902.7.1.1472.7.1.146 abAquifex14960750170819831390bcaeolicus3371554i1790 abThermotoga14691385020914150273amaritima0289 ebcAeropyrum20910768001217980011ipernix1109 ebPyrobaculum3437161008353055–iaerophilum0944 gebeArchaeoglobus–1494040123720108eicfulgidusVC1635614420230 fghbcPyrococcus09671199201301890049iabysii1515 fghbcPyrococcus03120196AF12790920141956ifuriosus0613 fghbcPyrococcus05891956164518970082ihorikoshii0759 ec,kHalobacteriumglcKgi10581425NP280577NP279697VNG0309CNRC-1 fhbcMethanococcus16041605160401090400bc09171585i0299 gbcMethanopyrus–NP614925–07411409ikandleri0954 fghbcMethanosarcinaNP6184430821NP61844433444591jcacetivorans11520439c3889c2666
ARCHAEA VOLUME 1, 2003
EMBDEN-MEYERHOF-PARNAS PATHWAY IN HYPERTHERMOPHILES 203
Table1continued.EnzymeCommission(EC)numbersandfour-digitidentificationnumbersordatabaseaccessioncodesofgenesidentifiedinhyperthermophilesandmesophilicarchaeathatencodeglycolyticen-zymes.Abbreviations:GLK=glucokinase;PGI=phosphoglucoisomerase;PFK=phosphofructokinase;FBPase=fructose-1,6-bisphosphatase;TPI=triosephosphateisomerase;GAP=glyceraldehyde-3-phos-phatedehydrogenase;PGK=phosphoglyceratekinase;iPGM=2,3-bisphosphoglycerate-independentphosphoglyceratemutase;dPGM=2,3-bisphosphoglycerate-dependentphosphoglyceratemutase;PK=pyruvatekinase;PPDK=pyruvate:orthophosphatedikinase;andPEPsyn=phosphoenolpyruvatesynthase. GLKPGIPFKFBPaseAldolaseTPIGAPPGKiPGM/EnolasePKPPDK,dPGMPEPsyn ECnumber(s)2.7.1.15.3.1.92.7.1.113.1.3.114.1.2.135.3.1.11.2.1.122.7.2.35.4.2.14.2.1.112.7.1.402.7.9.1,2.7.1.22.7.1.902.7.9.22.7.1.1472.7.1.146 bcdMethanobacterium–1546–0871057910411009104215910043–1118idΔthermoautotrophicumH16860418 bcdSulfolobus–2245–05472350203013561357377121216171235itokodaii03182120 bcdSulfolobus–2281–241832262592528527417091309810083isolfataricus02862236 iThermoplasma08251419–1428–0313110310751347088208960886acidophilum823d413 iThermoplasma07281438–1445–128704250494257098110200976volcanium730832d1158 abcdefgP.furiosusFamilyAPFKs;inositolmonophosphatasewithFBPaseactivity(MJ0109);DhnAaldolasefamily;iPGM;FamilyBATP-PFK;ADP-GLK(EC2.7.1.147);similartoPGI(additionallowerhijM.acetivoransThermococcoskodakaraensisscoringorthologsmaybePAB2301,PF0056,PH0047,MJ1618and2922);ADP-PFK(EC2.7.1.146);homologoustoKOD1(Rashidetal.2002);homologouskglpXM.jannaschiiP.abysiiP.furiosusP.horikoshii-encodedFBPase(Donahueetal.2000;lowscoringsequenceisMJ0865);ahighscoringparalogisVNG0683C;paralogousenolasesequencesfor,andareM.thermoautotrophicumA.fulgidusM.jannaschii0367,1641and1630,respectively(seetext);highscoringAF1425,MJ0010andMTH0418PGMsequencesaresuggestedtobephosphopentomutasesfor,andM.thermoautotrophicum,respectively(Galperinetal.1998);additionallowerscoringdPGMsequencesincludeTA0625,AQ1773,MA3008,TV1385andSSO2236(lattertwoidentifiedbyvanderOostetal.M.jannaschiiA.fulgidusA.fulgidusM.jannaschiiA.aeolicusM.thermoautotrophicum2002);gene0400and0108predictedtoencodealdolases(Dandekaretal.1999);gene1751,1612,1744and1591haveM.jannaschiibeenpredictedtoencodePGMs;multipleparalogoussequencesforFBPases,aldolases,PGMsandPEPsynthases/PPDKSareshownindecreasingorderofsimilarity(Evalue)usingtherelevantorT.maritimaSulfolobustokodaiiS.solfataricusgenes(TM1415/MJ0109forFBPases;TM0273/MJ1585foraldolases;TM1374/MJ1612forPGMs;TM0272/MJ0542forPEPsynthases/PPDKs).andeachcon-tainanopenreadingframeencodingonlytheN-terminalPPDKdomainofPEPsynthases/PPDKs(ST0698andSSOPpsA-2;Sonnhammeretal.1997).
ARCHAEA ONLINE at http://archaea.ws
204 RONIMUS AND MORGAN
Table2. Detectionofglycolyticenzymeactivitiesinhyperthermophilicprokaryotesandmesophilicarchaea.
Enzyme Species Reference
Glucokinase (ATP) Desulfurococcus amylolyticus Selig et al. 1997
Halorubrum saccharovorum Tomlinson et al. 1974
Sulfolobus solfataricus De Rosa et al. 1984
Thermoproteus tenax Siebers and Hensel 1993
Thermotoga maritima Schröder et al. 1994, Selig et al. 1997
Glucokinase (ADP) Archaeoglobus fulgidus7324 Labes and Schönheit 2001
Methanococcus jannaschii Sakuraba et al. 2002
Pyrococcus furiosus Kengen et al. 1994, 1995, 2001, Selig et al. 1997, Koga et al. 2000,
Verhees et al. 2002b
Thermococcus celer Selig et al. 1997
Thermococcus litoralis Selig et al. 1997, Koga et al. 2000, Ito et al. 2001
PGI (EC 5.3.1.9) Archaeoglobus fulgidus7324 Labes and Schönheit 2001
Desulfurococcus amylolyticus Selig et al. 1997
Haloarcula vallismortis Altekar and Rangaswamy 1992
Halobacterium halobium D’Souza and Altekar 1983
Haloferax mediterranei Altekar and Rangaswamy 1992
Methanococcus maripaludis Yu et al. 1994
Methanothrixstrain FE Pellerin et al. 1987
Pyrococcus furiosus Kengen et al.1994, Hansen and Schönheit 2001b, Verhees et al. 2001b
Sulfolobus solfataricus De Rosa et al. 1984
Thermococcus celer Selig et al. 1997
Thermoproteus tenax Siebers and Hensel 1993
Thermotoga maritima Schröder et al. 1994
PFK (Family A) Thermoproteus tenax(PPi) Siebers and Hensel 1993, 2001, Siebers et al. 1998
Thermotoga maritima(ATP) Schröder et al. 1994, Ding et al. 2001, Hansen et al. 2002
Thermotoga maritima(PPi) Ding et al. 2001
PFK (Family B) Aeropyrum pernix(ATP) Hansen and Schönheit 2001a, Ronimus and Morgan 2001
Desulfurococcus amylolyticus Ding 2000, Hansen and Schönheit 2000
PFK (Family C; ADP) Archaeoglobus fulgidus7324 Labes and Schönheit 2001
Methanococcusigneus Verhees et al. 2001a
Methanococcus jannaschii Verhees et al. 2001a, Sakuraba et al. 2002
Methanococcusmaripaludis Verhees et al. 2001a
Methanococcus thermolithotrophicus Verhees et al. 2001a
Methanosarcina mazei Verhees et al. 2001a
Pyrococcus furiosus Kengen et al. 1994, 2001, Tuininga et al. 1999
Thermococcus celer Selig et al. 1997
Thermococcus litoralis Selig et al. 1997, Ito et al. 2001
Thermococcus zilligii Ronimus et al. 1999, 2001a
FBPase Archaeoglobus fulgidus Stieglitz et al. 2002
Haloarcula vallismortis Altekar and Rangaswamy 1990, 1992
Halobacterium halobium D’Souza and Altekar 1983
Haloferax mediterranei Altekar and Rangaswamy 1992
Methanobacterium thermoautotrophicum Fuchs et al. 1983
Methanococcus jannaschii(MJ0109) Johnson et al. 2001
Methanococcus maripaludis Yu et al. 1994
Methanothrixstrain FE Pellerin et al. 1987
Pyrococcus furiosus Schäfer and Schönheit 1993, Verhees et al. 2002a
Thermococcus kodakaraensisKOD1 Rashid et al. 2002
Aldolase Archaeoglobus fulgidus7324 Labes and Schönheit 2001
Desulfurococcusamylolyticus Selig et al. 1997
Haloarculavallismortis Dhar and Altekar 1986, Krishnan and Altekar 1991
Altekar and Rangaswamy 1992
Halobacterium cutirubrum Dhar and Altekar 1986
Halobacterium halobium Dhar and Altekar 1986
Continued on facing page.
ARCHAEA VOLUME 1, 2003
EMBDEN-MEYERHOF-PARNAS PATHWAY IN HYPERTHERMOPHILES 205
Table2continued.Detectionofglycolyticenzymeactivitiesinhyperthermophilicprokaryotesandmesophilicarchaea.
Enzyme Species Reference
Adolase Halobacterium salinarum Dhar and Altekar 1986
Halobacterium volcanii Dhar and Altekar 1986
Halococcus saccharolyticus Johnsen et al. 2001
Haloferax mediterranei Dhar and Altekar 1986, Altekar and Rangaswamy 1992,
D’Souza and Altekar 1998
Halorubrum saccharovorum Dhar and Altekar 1986
Mb. thermoautotrophicum Fuchs et al. 1983
Methanococcus maripaludis Yu et al. 1994
Methanothrixstrain FE Pellerin et al. 1987
Pyrococcus furiosus Schäfer and Schönheit 1992, 1993, Kengen et al. 1994
Siebers et al. 2001
Thermococcus celer Schröder et al. 1994, Selig et al. 1997
Thermoproteus tenax Siebers and Hensel 1993, Siebers et al. 2001
Thermotoga maritima Schröder et al. 1994, Selig et al. 1997
TPI Archaeoglobus fulgidus7324 Labes and Schönheit 2001
Halococcus saccharolyticus Johnsen et al. 2001
Methanobacterium bryantii Hensel et al. 2001
Methanobacterium thermoautotrophicum Jansen et al. 1982, Fuchs et al. 1983
Methanococcus maripaludis Yu et al. 1994
Methanothermusfervidus Hensel et al. 2001, Schramm et al. 2001
Methanothrixstrain FE Pellerin et al. 1987
Pyrococcusfuriosus Schäfer and Schönheit 1992, Schäfer and Schönheit 1993
Pyrococcus woesei Hensel et al. 2001, Schramm et al. 2001
Thermococcus celer Selig et al. 1997
Thermoproteus tenax Siebers and Hensel 1993
Thermotogamaritima Schröder et al. 1994, Schurig et al. 1995b, Beaucamp et al. 1997,
Crowhurst et al. 2001
Thermotoga neapolitana Yu and Noll 2001
GAPDH Archaeoglobus fulgidus7324 Labes and Schönheit 2001
Desulfurococcusamylolyticus Selig et al. 1997
Haloarcula vallismortis Krishnan and Altekar 1991, Altekar and Rangaswamy 1992
Pruss et al. 1993
Halococcus saccharolyticus Johnsen et al. 2001
Haloferax mediterranei Altekar and Rangaswamy 1992
Halorubrum saccharovorum Tomlinson et al. 1974
Methanobacterium formicicum Fabry et al. 1989
Methanobacterium bryantii Littlechild and Isupov 2001
Methanobacterium thermoautotrophicum Zeikus et al. 1977, Fuchs and Stupperich 1982, Jansen et al. 1982
Methanococcus maripaludis Yu et al. 1994
Methanothrixstrain FE Pellerin et al. 1987
Methanothermus fervidus Fabry and Hensel 1987, Fabry et al. 1988
Pyrococcus furiosus Schäfer and Schönheit 1993
Pyrococcus woesei Zwickl et al. 1990
Sulfolobus solfataricus Littlechild and Isupov 2001
Thermococcus celer Selig et al. 1997
Thermoplasma acidophilum Budgen and Danson 1986
Thermoproteus tenax(GAPDH) Siebers and Hensel 1993
Thermoproteus tenax(GAPN) Brunner et al. 1998, Brunner and Hensel 2001
Thermotoga maritima Tomschy et al. 1993, Schröder et al. 1994, Selig et al. 1997
PGK Desulfurococcusamylolyticus Selig et al. 1997
Halococcus saccharolyticus Johnsen et al. 2001
Methanobacterium thermoautotrophicum Zeikus et al. 1977, Fuchs and Stupperich 1982, Jansen et al. 1982
Methanobacteriumbryantii Fabry et al. 1990, Hensel et al. 2001
Methanococcus maripaludis Yu et al. 1994
Methanothermus fervidus Fabry et al. 1990, Hess et al. 1995, Hensel et al. 2001
Methanothrixstrain FE Pellerin et al. 1987
Continued overleaf.
ARCHAEA ONLINE at http://archaea.ws
206 RONIMUS AND MORGAN
Table2continued.Detectionofglycolyticenzymeactivitiesinhyperthermophilicprokaryotesandmesophilicarchaea.
Enzyme Species Reference
PGK Pyrococcus furiosus Schäfer and Schönheit 1993, Hess et al. 1995
Pyrococcus woesei Hess et al. 1995, Hensel et al. 2001
Sulfolobussolfataricus Crowhurst et al. 2001
Thermococcus celer Selig et al. 1997
Thermoproteus tenax Siebers and Hensel 1993
Thermotoga maritima Schröder et al. 1994, Schurig et al. 1995b; Beaucamp et al. 1997,
Grattinger et al. 1998, Crowhurst et al. 2001, Hensel et al. 2001
Thermotoga neapolitana Yu and Noll 2001
PGM Archaeoglobus fulgidus7324 Labes and Schönheit 2001
Halococcus saccharolyticus Johnsen et al. 2001
Halorubrum saccharovorum Tomlinson et al. 1974
Methanobacterium thermoautotrophicum Zeikus et al. 1977, Fuchs and Stupperich 1982, Jansen et al. 1982
Methanococcus maripaludis Yu et al. 1994
Methanococcus jannaschii Graham et al. 2002, van der Oost et al. 2002
Methanothrixstrain FE Pellerin et al. 1987
Pyrococcus furiosus Schäfer and Schönheit 1993, van der Oost et al. 2002
Thermococcus celer Selig et al. 1997
Thermoproteus tenax Siebers and Hensel 1993
Thermotoga maritima Schröder et al. 1994
Enolase Archaeoglobus fulgidus7324 Labes and Schönheit 2001
Haloarcula vallismortis Altekar and Rangaswamy 1990, 1992
Halococcus saccharolyticus Altekar and Rangaswamy 1992, Johnsen et al. 2001
Haloferax mediterranei Altekar and Rangaswamy 1992
Halorubrum saccharovorum Tomlinson et al. 1974
Methanobacterium thermoautotrophicum Jansen et al. 1982
Methanococcus maripaludis Yu et al. 1994
Methanothrixstrain FE Pellerin et al. 1987
Pyrococcus furiosus Schäfer and Schönheit 1992, 1993, Peak et al. 1994
Thermococcus celer Selig et al. 1997
Thermoproteus tenax Siebers and Hensel 1993, Selig and Schönheit 1994
Thermotoga maritima Schröder et al. 1994, Schurig et al. 1995a, Selig et al. 1997
PK Archaeoglobus fulgidus7324 Labes and Schönheit 2001
Haloarcula vallismortis Altekar and Rangaswamy 1990, 1992
Halococcus saccharolyticus Johnsen et al. 2001
Haloferax mediterranei Altekar and Rangaswamy 1992
Halorubrum saccharovorum Tomlinson et al. 1994
Methanococcus maripaludis Simpson and Whitman 1993, Yu et al. 1994
Pyrococcus furiosus Schäfer and Schönheit 1992, Sakuraba et al. 1999
Thermoproteus tenax Siebers and Hensel 1993, Selig and Schönheit 1994
Schramm et al. 2000
Thermoplasma acidophilum Budgen and Danson 1986, Potter and Fothergill-Gilmore 1992
Thermococcus celer Selig et al. 1997
Thermotoga maritima Schröder et al. 1994
PEP synthase Aquifex pyrophilus Beh et al. 1993
Methanobacterium thermoautotrophicum Eyzaguirre et al. 1982, Fuchs and Stupperich 1982
Methanobrevibacter arborophilus Eyzaguirre et al. 1982
Methanococcus maripaludis Shieh and Whitman 1987
Methanothrixstrain FE Pellerin et al. 1987
Pyrococcus furiosus Schäfer and Schönheit 1993, Hutchins et al. 2001
Sakuraba et al. 2001
Staphylothermus marinus Cicicopol et al. 1999
FamilyBATP-PFKs(alsoEC2.7.1.11),whichcatalyzean Family B ATP-PFKs have been described for Aeropyrum
essentiallyirreversiblereaction,arerelatedtoFamilyBsugar pernix(HansenandSchönheit2001a,Ronimusetal.2001b)
kinasesthatcatalyzevarioussubstratephosphorylations,such and Desulfurococcus amylolyticus (Ding 2000, Hansen and
asthoseofadenosineandribose(Sigrelletal.1998).Archaeal Schönheit2000).OthersmaybepresentinPyrobaculumaer-
ARCHAEA VOLUME 1, 2003
EMBDEN-MEYERHOF-PARNAS PATHWAY IN HYPERTHERMOPHILES 207
ophilum(PAE0835;Fitz-Gibbonetal.2002),Archaeoglobus aeoglobus fulgidus (AF2372) and M. jannaschii (MJ0109)
fulgidus (AF0401, Cordwell 1999; or AF0356) and Halo- gene products support the idea of a close relationship with
bacteriumstrainNRC-1,asindicatedbyextentofsharediden- FBPases(Johnsonetal.2001,Stieglitzetal.2002).Versions
tityorsimilarity(Altschuletal.1997,RonimusandMorgan ofthisgenehavebeenfoundinM.thermoautotrophicumand
2001). Other high scoring sequences with the PFKB/ribo- Aquifex, Thermotoga, Aeropyrum, Archaeoglobus, Pyro-
kinasesignature(withEvalueslessthan10–7)arepresentin coccus and Sulfolobus species (Table 1). The FBPase from
allofthearchaealgenomes(Altschuletal.1997,Sonnhammer P.furiosushasrecentlybeencloned,expressedandcharacter-
et al. 1997; PEDANT: http://pedant.gsf.de). However, the ized(Verheesetal.2002a).Thisenzymewasmuchmorespe-
presence of ATP-PFK activity associated with any of these cificforfructose-1,6-bisphosphaterelativetoinositol-1-phos-
remains to be substantiated by biochemical data. phate(approximately13-fold)thantheMJ0109geneproduct
Adenosinediphosphate-dependentPFKs(ADP-PFKs;ten- (1.6-fold; Verhees et al. 2000a). It should be noted that the
tatively named Family C PFKs; EC 2.7.1.146) have been presenceofapyrophosphate-dependentPFKactivity(which
found in Thermococcuszilligii (Ronimus and Morgan 2001, makesthePFKreactionmorereversible)couldmakethere-
Ronimusetal.2001a),T.celer(Seligetal.1997),T.litoralis quirementforanFBPaseredundant(seebelow;Robersonet
(Selig et al. 1997, Koga et al. 2000), Pyrococcus furiosus al. 2001).
(Tuininga et al. 1999, Koga et al. 2000) and M. jannaschii ThediscoveryofasecondFBPaseinE.coli,encodedbythe
(Verheesetal.2001a),althoughthelastenzymeisapparently geneglpXandclassifiedasFBPaseII(Donahueetal.2000),
bifunctional, with ADP-dependent glucokinase activity and the FBPase in B. subtilis encoded by fbp (Class III
(Sakuraba etal.2002).It isalsopresentin somemesophilic FBPase), which shares no significant homology with other
methanogens (Table 2). The crystal structure of T. litoralis FBPases, has significantly affected the classification of
ADP-glucokinase (467 amino acids) has shown that the en- FBPases(Fujitaetal.1998).Followingrecentdescriptionsof
zyme has three-dimensional similarities to Family B sugar thearchaealFBPases/IMPasesandtheP.furiosusFBPase,a
kinases,includingthehumanadenosinekinaseandtheE.coli newclassificationschemehasbeenproposedwithfourclasses
ribokinase(eachapproximately33kDa),althoughthelevelof ofFBPasesandoneclassofIMPases(thelatterwithlittleorno
sequence identity is low (only seven strictly conserved resi- FBPaseactivity).TheClassIFBPases(classicalFBPasesof
duesinastructure-basedalignment;Itoetal.2001).Thelack bacteriaandeukarya),ClassIVFBPases(includingtheP.furi-
of sequence identity was such that a BLASTP comparison osusenzyme)andClassVIMPasessharesignificantsequence
withanexpectsizeof1000revealednosignificantsequence similarity(Verheesetal.2002a).Unfortunately,nostructures
similarity between the T. litoralis ADP-glucokinase and the areyetavailablefortheFBPaseClassIIorIIIenzymestosub-
E. coli ribokinase (Altschul et al. 1997; results not shown). stantiate potential distant sequence similarities.
Nevertheless, the authors proposed a common evolutionary AnotherrecentdevelopmentinFBPaseenzymologywasthe
origin for the two sequence families (Ito et al. 2001), and a cloning, expression, purification and characterization of an
FamilyBsugarkinasemotifwasfoundinasequence-based FBPase from the heterotroph Thermococcus kodakaraensis
comparison(Ronimusetal.2001b).Halococcussaccharolyt- KOD1(Rashidetal.2002).ThisFBPaseisspecificforfruc-
icus(Johnsenetal.2001),HaloferaxmediterraneiandHalo- tose-1,6-bisphosphate,incontrasttoFBPase/IMPases,andits
arculavallismortisutilizea1-phosphofructokinasetodegrade expression is repressed when strain KOD1 is grown in a
fructose,whereasH.saccharolyticus,H.saccharovorumand sugar-basedmedium.HighscoringorthologsoftheT.koda-
H.vallismortisuseamodifiedEntner-Doudoroffpathwayto karaensisFBPase(Evaluesrangingfrombetween10–152and
degrade glucose (Altekar and Rangaswamy 1992, Ranga- 10–117)arepresentinallofthearchaeainTable1exceptHalo-
swamyandAltekar1994).Thus,insummary,thecurrentdata bacteriumNRC-1andM.acetivorans.WhethertheFBPase/
(Tables1and2andabovediscussion)impliesthatthereisa IMPases(similartotheMJ0109geneproduct)ororthologsof
minimum of two sequence families to account for PFKs. theT.kodakaraensisFBPasefilltheroleofanFBPaseinother
archaealspeciesinvivoneedstobedeterminedbyadditional
Fructose-1,6-bisphosphatase biochemical evidence. However, their wide distribution in
Fructose-1,6-bisphosphatase(FBPase;EC3.1.3.11)catalyzes hyperthermophilesandarchaeaisintriguing.Thus,thecurrent
the conversion of fructose-1,6-bisphosphate to fructose-6- evidenceindicatesthattherearelikelytobefourseparatese-
phosphateingluconeogenesis,normallyabypassreactionof quencefamiliesencodingenzymeswithFBPaseactivityinthe
glycolysis.Aclassicalversionofthegene(i.e.,relatedtothat threedomains,withClassIVFBPasesandtheorthologsofthe
ofE.coliandhuman)isfoundonlyinHalobacteriumNRC-1 T.kodakaraensisenzymescommoninhyperthermophilicbac-
(Altschul et al. 1997) and not, as one might expect from its teria and archaea (Rashid et al. 2002, Verhees et al. 2002a).
widedistributionamongbacteria,ineitherAquifexorTherm-
Fructose-1,6-bisphosphate aldolase
otoga(http://pedant.gsf.de/).However,anenzymeinM.jan-
naschii(MJ0109)withinositolmonophosphataseactivity(EC Genes encoding aldolases (EC 4.1.2.13) that catalyze the
3.1.3.25),whichcatalyzesthedephosphorylationofinositol- reversible aldol cleavage of fructose-1,6-bisphosphate into
1-phosphate (IMP), has been shown to also possess FBPase glyceraldehyde-3-phosphateanddihydroxyacetonephosphate
activityandcouldactasanFBPase(IMPase/FBPase;Johnson arepresentintheThermotogaandAquifexgenomes(Table1;
et al. 2001). The IMPase/FBPase crystal structures of Arch- Cordwell1999,Dandekaretal.1999).Twoclassesofaldol-
ARCHAEA ONLINE at http://archaea.ws
208 RONIMUS AND MORGAN
asesaredistinguishableonthebasisoftheirreactionmecha- breakdownofglucose(Fothergill-GilmoreandMichels1993,
nisms(Fothergill-Gilmore andMichels1993):ClassIaldol- CopleyandBork2000).Theenzymeitselfhasbeendescribed
asesformaSchiff-baseintermediatebetweenthesubstrateand bysomeinvestigatorsascatalyticallyperfect because,based
alysineresidue,whereasClassIIaldolasesdonotformacova- on the K /K ratio, its reaction rate is limited only by sub-
cat m
lentintermediate.ClassIIenzymesarefoundinbacteriaand strate diffusion (Fothergill-Gilmore and Michels 1993).
lowereukaryotes(yeasts,fungiandalgae),whereasClassIen- Atotalof127TPIgeneswerefoundinthePfamdatabase
zymesarefoundinbacteria,protistsandmetazoa(Fothergill- (Sonnhammeretal.1997),andthesequencesaredistributed
Gilmore and Michels 1993). throughout the three domains of life (Cordwell 1999, Dand-
Recently,aClassIaldolasewasidentifiedinE.coli(dhnA ekaretal.1999,Koikeetal.1999).Atotalof31representative
gene product), and sequence analysis showed that orthologs sequencesfromthethreedomains,includingsequencesfrom
werepresentinseveralarchaealgenomesincludingArchaeo- archaeaandhyperthermophilicbacteria,wereusedtoproduce
globus,P.horikoshii,M.jannaschii,M.thermoautotrophicum an alignment and construct an associated phylogenetic tree
and Aeropyrum pernix, and also in the bacterium Aquifex (Figure1).WhenthesequencefromM.jannaschiiwasused
aeolicus (Dandekar et al. 1999, Galperin et al. 2000). Se- forsequenceretrievalfromtheTIGRComprehensiveMicro-
quenceanalysisshowsthatorthologstodhnAarealsopresent bial Database, % identities varied from 67% (Pyrococcus
inotherarchaea,excludingT.volcanium,T.acidophilumand abysii)to25%(BorreliabugdorferiB31)inananalysiscon-
P.aerophilum,andinsomecasestwoormoreparalogousse- taining TPI sequences from 66 genomes. The archaeal se-
quencesarepresent(Altschuletal.1997,Siebersetal.2001; quencesformaclearlyseparateddomain-specificclusterwith
see Table 1). aparticularlylongbranchlength,with100%bootstrapsupport
ThelackofanobviousaldolasegeneinP.aerophilumisin- from bothmaximumparsimonyandneighbor-joining analy-
teresting because this organism apparently has an otherwise ses.Theeukaryalsequencesgrouptogether,asdothebacterial
completeglycolyticpathway.Pyrobaculumaerophilumdoes, sequences,butthedomain-specificclustersarenotnearlyso
however,containaputativedeoxyribosealdolase(PAE1231) marked or bootstrap-supported as that of the Archaea.
thathasbeenshowntopossesssequencesimilaritytoDhnA- Triosephosphate isomerases of the hyperthermophiles
like aldolases and which could supply aldolase activity AquifexandThermotogadonotbranchasdeeplyasexpected
(Galperinetal.2000).Alternatively,aldolaseactivitymaybe onthebasisoftheir16SrDNAsequencecomparisons.How-
provided by an L-fuculose-1-phosphate aldolase encoded by ever, the Thermotoga TPI was isolated as a fusion complex
fucA,which,inE.coli,possessesupto4%fructose-1,6-bis- withbothTPIandphosphoglyceratekinaseactivities(Schurig
phosphate aldolase activity (Ghalambor and Heath 1966). et al. 1995b, Beaucamp et al. 1997, Hensel et al. 2001),
Morerecently,thealdolasesfromP.furiosusandT.tenaxhave whereas the usual TPI quaternary structure is homodimeric
beencloned,expressedandcharacterized(Siebersetal.2001). with no associated PGK activity (Fothergill-Gilmore and
These archaeal dhnA-like aldolases are also present in some Michels 1993, Hensel et al. 2001, Yu and Noll 2001). This
bacteria (including Aquifex and Thermotoga), but not in the couldaffecttherateofevolutionofboththeTPIandphospho-
Eukarya, and Siebers et al. (2001) have proposed the name glycerate enzyme sequences.
archaeal type Class I aldolase. For the archaeal TPIs, the crenarchaeal enzymes are the
Thoroughiterative-basedsequenceanalyseshaverevealed mostdeeplyrooted,whichisalsosomewhatunexpectedgiven
sequence signatures typical of classical Class II aldolases in thattheorganismsshowngrowaerobicallyand,basedon16S
ClassIdhnA-encodedarchaealaldolases,highlysuggestiveof rDNA sequence comparisons,are relatively late archaeal ar-
a distant relationship (Galperin et al. 2000). Furthermore, rivals (Woese et al. 1990). Furthermore, T. acidophilum and
basedontheircommonTIM-barrelstructuralfolds,anances- S. solfataricus do not process glucose via glycolysis but in-
traloriginfortheClassIandIIaldolaseshasbeenproposedas stead use the Entner-Doudoroff pathway (Selig et al. 1997,
part of a phylogenetic evolutionary history of central meta- Danson et al. 1998). Therefore, the evolutionary constraints
bolic TIM-barrel proteins (Copley and Bork 2000). In sum- operatingoncrenarchaeamayhavebeendifferentthanthose
mary,themostparsimoniousexplanationforthecurrentdata, thatexistedthroughouttheevolutionof,forexample,anaero-
althoughstillsomewhatcontroversial(Fothergill-Gilmoreand bic fermentative archaea. In general, TPIs are highly con-
Michels 1993), is for a single Schiff-base ancestral gene served and universally distributed, and a single sequence
havingevolvedintothepresent-dayrepresentativesofClassI familycanexplaincurrentlyidentifiedandcharacterizedTPIs.
and ultimately Class II aldolases after development of a
metal-dependentreactionmechanisminthelatter(Cooperet Glyceraldehyde-3-phosphate dehydrogenase
al. 1996, Galperin et al. 1998a, 2000). Glyceraldehyde-3-phosphatedehydrogenase(NAD+-depend-
ent GAPDH, EC 1.2.1.12; NADP+-dependent GAPDH, EC
Triosephosphate isomerases
1.2.1.13)isusuallyatetramerandcatalyzesthereversibleoxi-
Triosephosphateisomerase(TPI;EC5.3.1.1)isthearchetypi- dativephosphorylationofglyceraldehyde-3-phosphatetoform
cal (β/α) -barrel protein (TIM-barrel) that catalyzes the re- 1,3-bisphosphoglycerate (Forthergill-Gilmore and Michels
8
versibleinter-conversionofdihydroxyacetonephosphateand 1993).Thereactionmechanisminbacterialandeukaryoticen-
glyceraldehyde-3-phosphate, ensuring that, at least in gly- zymesinvolvesaconservedcysteineresidueintheactivesite,
colysis,twoATPmoleculesareeventuallyobtainedfromthe andtheenzymesfromthesetwodomainssharemorethan40%
ARCHAEA VOLUME 1, 2003