Hydrogenases from Methanogenic Archaea, Nickel, a Novel

33 Pages·2013·3.42 MB·English

ANRV413-BI79-18 ARI 27April2010 21:0
Hydrogenases from
Methanogenic Archaea,
Nickel, a Novel Cofactor,
and H Storage
2
g
or
s.
w
viey. Rudolf K. Thauer, Anne-Kristin Kaster,
ualree onl Meike Goenrich, Michael Schick,
ns
w.annal u Takeshi Hiromoto, and Seigo Shima
wo
ed from w3. For pers MemaaxilP:[e-mail],D-35043Marburg,Germany;
d1
wnloa06/10/ Annu.Rev.Biochem.2010.79:507–36 KeyWords
7-536. DoAustin on FMTihrasertcAhpnu1nb7ul,ias2lh0Re1de0voienwlinofeBaisocahRemevisiterwyiisnoAndlivnaenacteon rrHeesv2peaircrstaeitdvoaretylieoccnht,raeoinnn,etrcrghayen-mscfoeiornsvmerottiincgchoyudprliongge,nealseec,trcoonmbpilfeuxrcIaotfiotnh,e
79:50xas - biochem.annualreviews.org Abstract
10.Te Thisarticle’sdoi:
v. Biochem. 20y University of A01C00lol6.p1r61yi-gr44hi6g1t/h5sa4trn/e(cid:2)n1csue0rr2/ev00ve71.db00i7ob-cy0h5Ae0mn7n$.0u23a00l.05R00e8v.i1e5w2s1.03 aeaMcsnstoeoirvsctgaiytami-toecendotnhovafne[rNotHgiineF2gn,ei]ct-[hhNaeyryidcFrheuoa]sg-eeheanydrdaeisrfdeofuegrceeenonarCtseO[sN,2miFhweteeihtt]he-ahrnoHyoddp2irhsoutegolnfieandCzaeiHnsee4rs-.e,rdeFnudoacurmtcaitsenhelgye-
nu. Reb e[NneiFrgey]--hcoyndvroergteinngase[,NiaFned]-hFy4d2r0o-rgeednuacsiensgar[eNpiFhey]l-ohgyednreotgiceanllayser.elTathede
n
A to complex I of the respiratory chain. Under conditions of nickel
limitation, some methanogens synthesize a nickel-independent [Fe]-
hydrogenase (instead of F -reducing [NiFe]-hydrogenase) and by
420
that reduce their nickel requirement. The [Fe]-hydrogenase harbors
a unique iron-guanylylpyridinol cofactor (FeGP cofactor), in which
a low-spin iron is ligated by two CO, one C(O)CH -, one S-CH -,
2 2
and a sp2-hybridized pyridinol nitrogen. Ligation of the iron is thus
similar to that of the low-spin iron in the binuclear active-site metal
center of [NiFe]- and [FeFe]-hydrogenases. Putative genes for the
synthesis of the FeGP cofactor have been identified. The formation
ofmethanefrom4H andCO catalyzedbymethanogenicarchaeais
2 2
beingdiscussedasanefficientmeanstostoreH .
2
507
ANRV413-BI79-18 ARI 27April2010 21:0
Thenamehydrogenasewascoinedin1931by
Contents Stephenson&Stickland(2,3)foranactivityin
anaerobicallygrownEscherichiacolicellsmedi-
INTRODUCTION.................. 508
atingthereversiblereductionofdyeswithH .
H ASANINTERMEDIATEIN 2
2
DyereductionwasreversiblyinhibitedbyCO,
CH FORMATIONANDTHE
4
indicatingtheinvolvementofatransitionmetal
ORGANISMSINVOLVED....... 510
inH activation(4).Thetransitionmetallater
HYDROGENASESFOUND 2
turned out to be nickel in a binuclear nickel-
INMETHANOGENSAND
ironcenterinthecaseof[NiFe]-hydrogenases
THEIRFUNCTION............. 511
(5–8),ironinabinucleariron-ironcenterinthe
THEFOURSUBTYPES
case of [FeFe]-hydrogenases (9–11), and iron
OF[NiFe]-HYDROGENASES
in a mononuclear iron center in the case of
INMETHANOGENS............ 514
[Fe]-hydrogenase(12–14),whicharethethree
Energy-Converting
org [NiFe]-Hydrogenases........... 514 differenttypesofhydrogenasesknowntodate
ws. (Figure1)(15,16).
e Heterodisulfide
viy. From the work of Stephenson, it became
ualree onl R[NediFuec]t-aHsey-dArsosgoecniaatseedMvhADG.. 517 evidentthatmethaneformationfrombiomass
nnus in river sediments is at least in part the result
ww.aonal Me[tNhainFoep]-hHenydazroingee-nRaesdeuVchintgACG.. 520 of the syntrophic interaction of H2-forming
ded from w13. For pers CGoeen[NnezsiyFImnev]e-oHFlv4ye2dd0r-oiRngeednuacsiensgFrhABG.. 521 bmtuaercnttheeardinaooguseutnctshh.aAatsnindEte.inrsdcpoeleiecdiaenisndhlyHadte2rro-cgsoetnunsdutimerasi,nnsig-t
7-536. DownloaAustin on 06/10/ [FeN]-Hic[MkYNeaDiltFuRRereO]a-gtHGiuolyEnadtNi.ro.oAn.g.S.e..En..a..s..e.......................... 552232 t(fcdpeiayorHrnnbiasoimn<nai1acqcn0yuaacaPnelndaert;oikdEtbiaen(cid:4)it(sceHipvthiei+tcale/ybrHiteithmaaest)ops=fnoiasrsc−ttgtahe3ntehn0tae0Hptrmra2fololVccyroe)vnst(hse1cree7iynrn,m1tltor8hoaw)e--.
79:50xas - IWNITMHEOTHUTANCOYGTEONCSHROMES... 524 H2(2seethesidebartitle2dPropertiesofH2)(19,
2010.of Te StructuralProperties............... 525 2tr0o)n,ecvaernriaetrlobwetwcoenecneonrtrgaatnioisnms,sisbaencaiudseealitelceacn-
Rev. Biochem. by University H2CGSTaetnCOaeloysRftIaAinccGtvPooErrlovBVpeideIorAsitnyiCenFstHhe.G.e4.sPi.s....................... 552256 fEtisromsenteisslmyoaafdntHidefsf2uuaassrereeedthattnhroonautufugaaehlpllypcmyrfooteorxtmpihmlaeanasdtmoebgliyyecnmm1si5ec(0m1ro7mbo)r.rialgTlnaioehnsne-.
nnu. FORMATION.................... 527 combustionof150milliontonsH2yields18 ×
A 1018 J,anenergyamountthatis3.75%ofthe
primaryenergyconsumedin2006bytheworld
INTRODUCTION
population(455 × 1018J).
In 1933, Stephenson & Stickland (1) en- Today on Earth, most of the H used
2
Blackandwhite richedfromriversedimentsmethane-forming by methanogens is of biological origin. Only
slimkeoskterursc:tucrhesimfonremy-ed microorganisms that grow on H2 and CO2 some of the H2 that sustains the growth of
(Reaction 1) and concluded that these methanogensisgeochemicallygenerated,e.g.,
aroundhydrothermal
vents,where methanogens must contain hydrogenases that inblackandwhitesmokers.However,intheAr-
superheatedmineral activateH2(Reaction2). chaeozoic(4to2.5billionyearsback),whenthe
richwaterfrombelow 4H +CO →CH +2H O differentlineagesofmicrobesonEarthevolved
Earth’scrustcomes 2 2 4 2
andwhenthetemperaturesweremuchhigher
throughtheocean (cid:2)G◦(cid:4)=−131kJmol−1. 1.
than today, geochemically formed H proba-
floor 2
H (cid:2)2e−+2H+ E(cid:4) =−414mV. 2. blypredominatedthatofbiologicaloriginand
2 o
508 Thaueretal.
ANRV413-BI79-18 ARI 27April2010 21:0
Large
Open
subunit Cys N
site C Cys
S
Ni S Fe CN Cys S
Cys S C SS Fe
S Cys 6 Å Cys S Cys O Cys Fe S
Fe S S Cys Open X S S FeS SFe
site 4Å
S Fe S
Fe Fe S
S Fe NC CN
Fe SSCys Small OC OC CO Cys
Cys S subunit
[NiFe]-hydrogenase [FeFe]-hydrogenase
Cys
g O
or S
ws. OC
e Fe
viy. N
nualrese onl Ositpeen OC HO OGMP
nu
ww.aonal [Fe]-hydrogenase
m wpers
ed fro3. For FTihgeumree1talsitesofthethreetypesofhydrogenasesinvolvedininterspecieshydrogentransfer(seeFigure2)
7-536. DownloadAustin on 06/10/1 hhrbeiayolvdasertyeoundgntehauntesasutishaseels(s1l(te25rv)–ue.8clAt)ou,bf[rbFatrhleefeFveieiraa]ttp-uihroriyenmds:raGionrygMcesonPtmra,usgmceutsoua(nnr9e,y–lso1yu1rlc)rah,etasatntsh.dien[ltFerveine]-lshiocyfdCtrhOoegleeinngazanysemds(e1.s2Di–ne1vs4po)iltvaeeredthniinsotftahpcehti,yr[laoNcgtieFivneee]-t-siictaelly
10.79:50Texas - fsuisetleendtlyt,heamgoronwgthreocefntmehthyadnrooggeennos.troCpohnic- ibneeBnafcotuernidaiannAdrlcohwaeear(E15u,k1ar6y)a., have not yet
20of methanogens, there are many hyperther-
em. sity mophiles,suchasMethanopyruskandleri(98◦C
BiochUniver odopctoicmcuusmjangnroaswchthiit(e8m5◦pCeroaptutirme)uamndgMroewththantoecmal-- PROPERTIESOFH2
Rev. by perature),andthesehyperthermophilesbranch H2isacolorlessgaswithaboilingpointat22.28K(−250.87◦C).
u. offthe16Sphylogenetictreerelativelyearly. Its Bunsen coefficient α in water at 20◦C is 0.018 (0.8 mM at
n
An This review highlights the properties of 1bar),anditsdiffusioncoefficientDw inwaterat20◦Cisnear
the five different hydrogenases found in 4 × 10−9m2s−1.ThehomolyticcleavageofH inthegasphase
2
methanogenswithinthecontextoftheirfunc- isendergonicby+436kJmol−1,andtheheterolyticcleavagein
tion in metabolism. Four of the enzymes waterat20◦Cisendergonicbyabout+200kJmol−1 (pK near
a
are[NiFe]-hydrogenaseswithsomeproperties 35) (19). The combustion energy of H is 120 MJ kg−1. The
2
similar and others dissimilar to those of re- activationofH ismechanisticallychallenging,andthecatalytic
2
lated [NiFe]-hydrogenases in bacteria. It was mechanism is of considerable interest. The H generated, e.g.,
2
in methanogens that nickel was first found to byelectrolysisorphotolysisofwater,ispresentlydiscussedasan
be required for hydrogenase activity (21, 22). environmentallycleanenergycarrierforuseinfuelcell-powered
The fifth enzyme is a [Fe]-hydrogenase (23) electricalcars.BeforeH canbeusedinfuelcells,cheapcatalysts
2
that is unique to methanogens and functional still have to be developed, and it is hoped that the active-site
in these only under conditions of nickel limi- structureofhydrogenaseswillshowhowtoproceed(20).
tation.[FeFe]-hydrogenases,whicharepresent
www.annualreviews.org • HydrogenasesfromMethanogens 509
ANRV413-BI79-18 ARI 27April2010 21:0
Anoxic environments CH4 Atmosphere
Freshwater and marine sediments, (1.8 ppm)
wetlands, swamps, intestinal
tracts of ruminants and termites, 0.4 Gt per year
hot springs, hydrothermal vents
Methanogenic archaea
with cytochromes Oxic
[NiFe] environments
Acetate- Aerobic
+ H+ bacteria
Rate- CO2
limiting Propionate 0.6 Gt per year
step Butyrate Acetogenic
bacteria CO2 + CH4
Biomass Monomers Lactate CO2
[FeFe] and [NiFe] ~1 Gt per year
(~2% NPP) Ethanol Anaerobic
H2 + CO2 archaea and
(pH2 < 10 Pa) bacteria
Bacteria [FeFe] and [NiFe]
g Protozoa [FeFe] H2 + CO2
or
s. Fungi [FeFe]
viewy. Archaea [NiFe] Mwieththoaunto cgyetoncich arorcmheasea
w.annualrenal use onl RDeiffauctsiioonns GeochHe2[mNiiFcea]l and [Fe] (>M10e,t0h0a0n Ge th mydertahtaense)
wo
m wpers
ed fro3. For FAipgpurroexi2mately2%ofthenetprimaryproduction(NPP)ofplants,algae,andcyanobacteriaarefermentedin
79:507-536. Downloadxas - Austin on 06/10/1 saraippantubnnoravmobodttoeperxiclneiinvHacovteatinieanas2aanblttictesnvooeo,ditlf,rnimeoctb[rcNhneuseseptumtnihyeFbHtaercresanniat+]etrt,otes/ias[goH,tFHbenelaey2ns2cFr.sacetteoacmrs]toauy,eannnpa,isvnltnaefredensorardbpt[.tiFehnTepleitgoHch]h,waeaarc7nsek0seosiio.tnsp1laceetcniμeatcoeiMttncatiioosvrcene(a−m<lnnyo3ted1fr(0t0ashat0ethnaPiemeonaareFne)Vmrsi(ao.g1obnbTu7dudir,hcyielC1nedm8a1Otui)mh)c..p2rrAiIeocaanetsronettrotdhhygflepaecatscanehieknsniesiltonmbopefgewsrsh.touiwTyHcnsdeieaht2srldhesoctrbgrmaoearyenfceenoctttrahsehoeusneae,fcntshahrteecaoi,ntretshiimtvtnoaaoanittaltsesvet,p,ahestrndeahoanasecbtdreoeresealsiddctoyh-xat
10.Te
20of
Biochem. University HCOHR2 AG4SFAOANNRISMIMNASTTIIENORVNMOAELNDVDIEADTTEHEIN dexecgIrrneatdeeaddbrbayyteae-nlxiatmeraricotiebnliglculbasrtaechptey,rditrahoelayntdibciopemrnozatysosmzoeiass
Rev. by Approximately2%ofthenetprimaryproduc- tomonomers,whichafteruptakebythesemi-
u. tionofplants,algae,andcyanobacteria(70bil- croorganismsareprimarilyfermentedtolactic
Ann lion tons C per year) are remineralized via acid,propionicacid,butyricacid,ethanol,and
aceticacidwiththeconcomitantformationof
methaneinanoxicenvironmentssuchasfresh-
CO ,formicacid,andsomeH .Thisprocess
waterandmarinesediments,wetlands,swamps, 2 2
alsoinvolvesanaerobicfungiintherumenand
sewage digesters, landfills, hot springs, and
anaerobicarchaeainhotsprings.Oftheseprod-
the intestinal tract of ruminants and termites
ucts, lactic, propionic, and butyric acids and
(Figure2).Fromthebiomass,whichconsistsof
ethanolservesyntrophicbacteriaassubstrates,
60%–70%cellulose,approximatelyonebillion
Geochemically
which ferment them to acetic acid, CO , H ,
formedH2: H2 tonsofmethanearegeneratedperyear;60%is andformicacid. 2 2
generatedabiotically oxidizedtoCO bymicroorganisms,and40%
2
fromH2Sorby escapesintotheatmosphere,whereitsconcen- From acetic acid, H2, CO2, and formic
reactionofH2Owith trationalmostdoubledwithinthelasthundred acid,methaneisthenformedbymethanogenic
ultramaficrocks archaea, of which there are two types, those
years(17).Thisisofconcernsincemethaneis
(serpentinization)
with and those without cytochromes. Acetic
aneffectivegreenhousegas.
510 Thaueretal.
ANRV413-BI79-18 ARI 27April2010 21:0
acidisconvertedtoCO andmethaneonlyby of glucose from cellulose to three CO and
2 2
the methanogens with cytochromes, and H , three CH involves 12 H as intermediates,
2 4 2
CO and formate are converted to methane which underlines the quantitative importance
2
mainly by those without cytochromes. None of H as electron carrier between fermenters
2
of the methanogens can use lactic, propionic, andmethanogens.
or butyric acid as energy substrates. But by All methanogens are known to belong to
consuming H , acetic acid and formic acid, the domain of Archaea and to the kingdom
2
themethanogenskeeptheH partialpressure ofEuryarchaeota.Fromthelatterlineage,the
2
between 1 Pa and 10 Pa and the acetic and Methanopyrales branch off first, followed by
formicacidconcentrationswellbelow0.1mM, theordersMethanococcalesandMethanobac-
enabling the syntrophic bacteria to convert teriales, and then by Methanomicrobiales and
lactic,propionic,andbutyricacidandethanol Methanosarcinales. Only the members of the
to acetic acid, H , and CO . Only at low Methanosarcinales contain cytochromes and
2 2
org concentrations of H2 and acetic acid are the can use acetic acid as methanogenic sub-
ws. fermentations of the syntrophs exergonic strate. Methanogenesis from acetate is there-
e
viy. enoughtosustaintheirgrowth(18). forebelievedtobealateinvention.Theabil-
ualree onl Intheintestinaltractofruminantsandter- ity to use acetate as methanogenic substrate
nnus mites,methanogenswithcytochromesarenot was associated with a change in the mecha-
w.anal present,andthereforemethanogenesisfromac- nismofenergyconservation,aselectrontrans-
wo
m wpers etate does not occur. The reason for this is port now involves cytochromes. The altered
ed fro3. For pmreotbhaabnloygtehnastisthgeengerroawlltyhlorawteerotfhaancetthoecldaisltuic- mtoecchhroanmisemstoalalolswoeudsethmeemtheatnhoaln,omgeetnhsywlaimthinceys-,
d1
wnloa06/10/ ttihoenarcaetteocinlastthiceminettehstainnoagletrnascat,reancdontthinerueofuosrley, aanlsdo mhaedthaylpthriicoels, naasmeenleyrgtyhesulobssstroatfetsh.eBaubtili-t
7-536. DoAustin on weacsaiissdhfbreoudmiolduastc.ueBptiecccoaancuissdiedtoehfreathbceloynla(c>cekn1ot0rfammtiMeotnh)oawnfitoahcgeetthniec- ilwtoyhwictoh10iussPaeacHh(af2roardcotaewnrnisetitxcopolpafnamratteiiatohlna,pnroseegseseunrbseewsloibtwhe)--,
79:50xas - resultthat,forthethermodynamicreasonsdis- outcytochromesthatarespecializedonH2plus
10.Te cussedabove,lactic,propionic,andbutyricacid, CO2 and/orformateasenergysources.Mem-
20of thereforealsoincreaseintheirconcentrations. bers of the Methanosarcinales that can grow
v. Biochem. y University Tnusahenedtsfoaornrgdagnilnuicsceoacnctseidofrsgoeamnreetshriseeis(rolairncbtteiecdstaibnnydaltpthrraeocprtisuoamnniidc- oohnfigthHheer2oHatnh2dercCoonOrcd2eenrdtsor,awttihohinsicshontlhlayacnkattchyseitgonmchifiermcoambneetlrsys.
Reb acid)andATPsynthesis(aceticacidandbutyric ThisiswhytheMethanosarcinalesdonotcon-
nnu. acid). tribute to methane formation from H2 and
A In sediments of hot springs, in which cel- CO in most anoxic environments (Figure 2)
2
luloseiscompletelyconvertedtomethaneand (17).
CO ,surprisingly,attemperaturesabove60◦C,
2
acetoclasticmethanogensareabsentforreasons
HYDROGENASESFOUND
notyetfullyunderstood.Methanogenswithcy-
tochromesgrowingabove60◦Chaveyettobe INMETHANOGENSAND
THEIRFUNCTION
found.Inhotsediments,aceticacidisconverted
to two CO and four H by bacteria related The genomes of several members of each
2 2
to acetogenic bacteria, and the H and CO of the five known orders of methanogens
2 2
thus formed are then converted to methane have been sequenced, and the genes puta-
by methanogens without cytochromes, which tivelyencodinghydrogenaseshavebeeniden-
havemanythermophilicandhypothermophilic tified. Biochemical studies of the hydroge-
species. Thus, in hot springs, the conversion nases have concentrated mainly on a few
www.annualreviews.org • HydrogenasesfromMethanogens 511
ANRV413-BI79-18 ARI 27April2010 21:0
species, namely Methanothermobacter thermau- of methanogens without cytochromes (maxi-
totrophicus, Methanothermobacter marburgensis, mally 3 g per mole CH ), indicating that the
4
Methanococcusmaripaludis,Methanosarcinabark- ATP gain per mole methane is approximately
EchA-F,EhaA-T,
eri, and Methanosarcina mazei. Genetic analy- 0.5 in methanogens with cytochromes and
EhbA-Q,and
MbhA-N: energy- seshavebeenrestrictedtoMethanococcusvoltae, 1to1.5inmethanogenswithoutcytochromes
converting[NiFe]- M. maripaludis, Methanosarcina acetivorans, (17). The low ATP gain of 0.5 allows the
hydrogenases M.mazei,andM.barkeri.Fromthesestudies,a methanogenswithoutcytochromestogrowon
MvhADG: partiallycoherentpicturehasemerged. H and CO at H partial pressures of 5 Pa
2 2 2
heterodisulfide Four different subtypes of [NiFe]- atwhichmethanogenesisfromCO andH is
2 2
reductase-associated hydrogenases and one [Fe]-hydrogenase exergonicby−25kJpermole,whichisjustsuf-
[NiFe]-hydrogenase
are found in methanogens. The four [NiFe]- ficienttodrivethesynthesisof0.5moleATP
HdrABCand subtypes are (a) the membrane-associated, ((cid:2)G(cid:4) = −50 kJ per mole ATP). Conversely,
HdrDE:
energy-converting [NiFe]-hydrogenases anATPgainof1to1.5isonlythermodynami-
heterodisulfide
org reductases (EchA-F, EhaA-T, EhbA-Q, and MbhA-N) callypossibleiftheH2concentrationis>100Pa
ws. VhtACG: for the reduction of ferredoxin with H2; ((cid:2)G < −63 kJ per mole CH4). And indeed,
e
viy. methanophenazine- (b) the cytoplasmic [NiFe]-hydrogenase methanogens with cytochromes are known to
ualree onl reducing[NiFe]- (MvhADG) associated with the heterodisul- have a much higher H2 threshold concentra-
nnus hydrogenase fide reductase (HdrABC) for the coupled tion(>100Pa)thanmethanogenswithoutcy-
ww.aonal FrhABG: anF420- reduction of ferredoxin and of the het- tochromes <10 Pa) (17). The relatively high
ed from w3. For pers rheyddurcoignegn[aNseiFe]- emrereodmduicbsiunralgfindee-a[NsCsoioFcMiea]-t-Sehd-yS,d-rCoogmeBneawtsheitahnoH(pVh2he;tnA(ac)CzinGthe)e-; methrearemllsyhbeonrlodstcooifnnvtchoeenlvterMdateiiotnhnafmnoorestHahra2cnicnoaangleeensxepaslriaseinfgrweohnmy-
d1
wnloa06/10/ raendduci(ndg) th[NeiFcey]t-ohpyldasrmogiecnacsoeesnzy(FmrehAFB4G20)-. H(F2igaunrde2C)Oan2dinwhmyoisntsmometehamnoemgebneircs,hea.bgi.t,aitns
Doon Not all five hydrogenases are found in all M.acetivorans,transcriptionofthegenesforthe
7-536. Austin mreedtuhcainnogg[eNnsi.FeT]-hhuysd,rothgeenmaseethiasnroepshtreinctaezdinteo- hydFroorgeannaseusnadreersptearnmdianngenotflytthuernfeudncotfifo(n24o).f
79:50xas - methanogens with cytochromes, and the thedifferenthydrogenasesintheproposedtwo
10.Te cytoplasmic[Fe]-hydrogenase,whichtogether metabolicschemes(Figure3a,b)theenergetics
20of with F -dependent methylenetetrahy- offerredoxinreductionwithH areofspecial
em. sity dromethan42o0pterin dehydrogenase substitute importance.Underphysiologica2lstandardcon-
v. Biochy Univer fuonrderthe nFi4c2k0e-lr-eldimucitiinngg [NgiFroew]-thhydrogcoennadsie- tdhiteiornesdu(pcHtio2n=of1f0e5rrPead;opxHin7(;EF(cid:4)odo=x/F−d4re2d0=m1V)),
nu. Reb tmioenthsan(soegeenbselowwi)t,hoiustonclyytopchrersoemnets.inGseonmees wgoitnhicHn2or(Ee(cid:4)oxe=rgo−ni4c1.4HmowVe)veisr,nuenitdheerrienndvievro-
n
A for [Fe]-hydrogenase synthesis are lacking in conditions(pH = 10Pa;pH7;Fd /Fd <
2 ox red
methanogens with cytochromes and in most 0.01), the reduction of ferredoxin with H is
2
membersoftheMethanomicrobiales(15,23). strongly endergonic with E(cid:4) of the H+/H
2
The function of the different [NiFe]- couple = −300mVandthatoftheFd /Fd
ox red
hydrogenasesinmethanogenesisfromH and couple = −500mV.Inmethanogens,fullyre-
2
CO inmethanogenswithcytochromescanbe ducedferredoxinisrequiredforthereduction
2
deducedfromFigure 3aandinmethanogens of CO to formylmethanofuran (CHO-MFR)
2
withoutcytochromesfromFigure3b.Thedif- (E(cid:4) = −500 mV), which is the first step in
o
ferencesoutlinedinthetwoschemesarebased, methanogenesisfromCO (Figure3),forthe
2
amongmanyotherobservations,onthefinding reduction of CO to CO (E(cid:4) = −520 mV),
2 o
thatthegrowthyieldofcytochrome-containing forthereductionofacetylcoenzymeA(acetyl-
methanogensonH andCO (maximally6.4g CoA)andCO topyruvate(E(cid:4) = −500mV),
2 2 2 o
permoleCH )ismorethantwiceashighasthat and—inmostmethanogens—forthereduction
4
512 Thaueretal.
ANRV413-BI79-18 ARI 27April2010 21:0
a b
Biosynthesis Biosynthesis
CO CO
2 2
2 H+ [NiFe]Fdred2– 2 Na+ [NiFe] Fdred2–
EchA-F H2 + Fdox CHO-MFR EhaA-T H2 + Fdox CHO-MFR
and/or
EhbA-Q
+ – H2F[rNhiAFeB]G F420HC2H HM4MtdPT+ + – H2F[rNhiAFeB]G F420HC2H HM4MtdPT+
H2F[rNhiAFeB]G F420HC2H2=H4MPT 2N aH++ H2F[rNhiAFeB]G F420HC2H2=H4MPT 2N aH++
VhtACG
[2N HiF+e] CH3-H4MPT CH3-H4MPT
nualreviews.orgse only. 2 HH+2 HdMP MPHr2DE HSC-oCMo+M- S2 - +HS -+HCSo-BCoCBHC3-HSN4-iCoAATMDPP 24 NH+a+ 2 H2[NiFFe]dCooxMMHvF-dhSdrA-ArSHeHDB-dSSCGC2--–C/oCoBoMBCHC3-HSN4-iCoAAMTDPP 24 NNaa++
nu
ww.aonal Reaction Diffusion
ed from w3. For pers FTihgeuprero3posedfunctionandlocalizationwithinthecellofthe[NiFe]-hydrogenasesinvolvedin
wnload06/10/1 mcmyeatottthceahrnrooofmgdeeinssep.suTisthef,eroacnmadtiHothn2esastnrtoadnicCshlOoioc2mataeerdterysahnoodfwethnleefcoetrrxoamnctebtsihtfouainrccohagitoeiomnnse(itanry)twhoefitthMhcevyhttrAoacDnhsGrloo/cmHatedisoranAnBrdeCa(bcc)toiwomnitpshlaoeruxetrsetmillaains
Doon tobeascertained(17).InsomemembersoftheMethanomicrobiales,genesforMvhAandMvhGarenot
7-536. Austin foofutnhdes,eanndoveenlefirgnyd-icnogns,vseeretinthgehsyedcrtioognenHaesetesrootdhiesurltfihdaenREehdauocrtaEseh-bAcssaoncbiaetepdre[sNeniFte(]2-5H).yFdororginentearsperetations
em. 2010.79:50sity of Texas - mMrtmraaetevhntthhhesAfyreonDlttrhyemGlattrane,attbhitroeeyantlhdro.ryawAohd.mrybModbemrrteoehmevmatinbhaeeoattirnhposoatneonpsrfot:ienptCrh;tiHeenCr;MOionCB-e(HM-tHSh2FHa4==nMRH,o,cPs4foaoTMerrcn)mPi.znTTyyaml,lhmemeeseetBatwthnhwaodynilpMtoehtnfeeuierttrtihsaneanttshnr;haioCohaclvHyoegdc≡rrcioodaHumelenp4seM;tticChcoPaaonlTnMtfoa+uip-nn,StceHttreiiot,nrnc;aoshCeiynnHdzrC3yo-m1sH-aeur4cnMMiintPawpTitt,ehriitns
BiochUniver thhyidorloggreonuaps;eCs;oFMd,-fSe-rSre-CdooxBin,hweittehrotwdiosu[l4fiFdee4;SE]-cchlAus-tFe,rEs;hFarAh-ATB,Gan,dFE42h0b-rAe-dQuc,iennger[Ngyi-Fceo]n-hveyrdtrinogge[nNaisFe;e]-
v. y HdrABCandHdrDE,heterodisulfidereductases;MP,methanophenazine;MvhADG,heterodisulfide
u. Reb reductase-associated[NiFe]-hydrogenase;VhtACG,methanophenazine-reducing[NiFe]-hydrogenase;
n VhtCandHdrE,b-typecytochromes.
n
A
of succinyl-CoA and CO to 2-oxoglutarate complex,themechanismofcouplingisbyelec-
2
(E(cid:4) = −500mV).Inmethanogensgrowingon tronbifurcation(Figure3b).
o
H and CO , the latter three reduction reac- Whereas the reduction of ferredoxin with
2 2
tions participate in autotrophic CO fixation. H inmethanogensisstronglyendergonic,that
2 2
Alloftheseferredoxin-dependentoxidoreduc- of methanophenazine (E(cid:4) = −170 mV) and
o
tase reactions are catalyzed by cytoplasmic of the heterodisulfide CoM-S-S-CoB (E(cid:4) =
o Electronbifurcation:
enzymes.Therefore,itisthereductionofferre- −140 mV) with H2 (E(cid:4)o = −414 mV) is a thedisproportionation
doxinwithH thatmustbeenergydrivenand stronglyexergonicreaction(26).Consistently, oftwoelectronsatthe
2
thesiteofenergycoupling.Asoutlinedbelow, methanophenazineandCoM-S-S-CoBreduc- sameredoxpotential
tooneelectronwitha
inenergy-converting[NiFe]-hydrogenases,the tion with H are coupled with energy con-
2 higherandonewitha
mechanismofenergycouplingischemiosmotic servation (Figure 3a,b). Of the hydrogenase-
lowerredoxpotential
(Figure 3a,b),andintheMvhADG/HdrABC catalyzed reactions, only the reduction of
www.annualreviews.org • HydrogenasesfromMethanogens 513
ANRV413-BI79-18 ARI 27April2010 21:0
coenzymeF (E(cid:4) = −360mV)withH isnot between the two subunits and close to the
420 o 2
coupled with energy conversion (27). Under proximal [4Fe4S]-cluster of the small subunit
invivoconditions(pH = 10Pa;F /F H (Figure 1a). A gas channel connects the sur-
2 420 420 2
<0.1), the free energy change associated with facewiththeactivesite(30).
the reaction is essentially zero. The energetic The large subunit of most [NiFe]-
differencesofthehydrogenase-catalyzedreac- hydrogenases is synthesized as a preprotein.
tionsinmethanogenesisfromH andCO can The C-terminal extension after H/R of the
2 2
therefore explain why there are at least three DPCxxCxxH/Rmotifisclippedoffproteolyt-
different hydrogenases in hydrogenotrophic ically in the maturation process (31–33). The
methanogens. gene coding the large subunit of some of the
energy-converting hydrogenases (34, 35) and
some of the H -sensory [NiFe]-hydrogenases
2
THEFOURSUBTYPES
(36) ends with a stop codon directly after the
ws.org OINFM[NEiTFeH]-AHNYODGREONGSENASES nmuoctliefo.tTidheerseefqoureen,cseynftohrestihseofDtPhCesxexC[NxxiHFe/]R-
e
viy. The crystal structures of the [NiFe]- hydrogenases appears to be independent of
ualree onl hydrogenases found in methanogens have not thisproteolyticmaturationstep.
nnus been determined. Currently, only structures Inthenextparagraphs,wesummarizewhat
w.anal of[NiFe]-hydrogenasesfromsulfate-reducing is known about the four different subtypes of
wo
m wpers bacteria are available (5–8). However, on the [NiFe]-hydrogenases found in methanogens:
ed fro3. For bhaysdirsogoefnasseeqsueanpcpeeacromtopabreisopnhsy,loagllen[eNtiicFaell]y- ((ab))heetneerrogdyi-sucolfindveerrteidnugcta[Nse-iFases]o-hciyadterodg[NeniaFsee]s-,
d1
wnloa06/10/ riselastoedm,etiamltehsougrehstritchteedsetqouetnhcee sseiqmuielanrcietys h[NydiFroeg]-ehnyadsreo,g(ec)namsee,thaanndoph(de)nazFi4n2e0--rreedduucciinngg
Doon around the N-terminal and C-terminal CxxC [NiFe]-hydrogenase.
7-536. Austin mreosptiefcst,ivRelxyC, GinxvColxvxexdHinan[NdiFDeP]-CcexnxtCerxxcHo/oRr-, Energy-Converting
79:50xas - dination.Nevertheless,itisgenerallyassumed [NiFe]-Hydrogenases
10.Te that the active-site structures of all [NiFe]- Energy-converting[NiFe]-hydrogenasesfrom
20of hydrogenases are very similar (Figure 1a);
v. Biochem. y University bfervuoitdmeinnRcaoelnstteohnaciataseteuhte(rsoopllihugaba)nl,edth[NestrreiuFicest]u-shpreeycdctrrooougslecdnoapbsieec dma(Elryei(cid:4)zvt≈heenant−bhoye5g0ear0npesvmraoerVrteso)imnbwleoeimrthrsbeorHddauni2ucemt(iaEos(cid:4)insoo=ncoimaf−toef3edtir0var0eneddfmoocVrxacit)ne-,
nu. Reb sub[sNtainFteia]-llhyyddirfofegreennats(e2s8,ar2e9).minimally com- (Reaction3)(Figure4)(34,35).
An posedoftwosubunits,alargeone(40–68kDa) Fd +H +(cid:2)μH+/Na+(cid:2)Fd 2− +2H+.
ox 2 red
and a small one (16–30 kDa). The large sub- 3.
unit harbors the [NiFe]-binuclear active-site Related enzymes are found in some
center. The small subunit generally contains hydrogenotrophic bacteria and in some
threelinearlyarrangedandevenlyspacediron- H -forming bacteria and archaea. In the H -
2 2
sulfurclusters,aproximalandadistal[4Fe4S]- formingmicroorganisms,theenzymecatalyzes
cluster,andonecentral[3Fe4S]-cluster(8).In the reverse of Reaction 3. Most convincing is
energy-converting [NiFe]-hydrogenases, the theenergy-convertingfunctionofthe[NiFe]-
small subunit contains only the proximal hydrogenase in the gram-negative Rhodospiril-
[4Fe4S]-cluster,whichappearstobenecessary lumrubrum(37)andRubrivivaxgelatinosus(38)
and sufficient for [NiFe]-hydrogenase func- aswellasinthegram-postiveCarboxidothermus
tion. In the heterodimer, the [NiFe]-center is hydrogenoformans (39). These anaerobic bac-
buriedandlocatedclosetothelargeinterface teria can grow chemolithoautotrophically on
514 Thaueretal.
ANRV413-BI79-18 ARI 27April2010 21:0
CO,withH andCO beingtheonlycatabolic
2 2
endproductsformed(CO+H O(cid:2)CO +H
2 2 2
(cid:2)G◦(cid:4) = −20 kJ mol−1) (40). The fermen-
H2
tation, which involves only a cytoplasmic
31–69 1x
carbon monoxide dehydrogenase (CooS), a kDa [4Fe4S] [NiFe] E' = –300 mV
cytoplasmic polyferredoxin (electron transfer ΔμH+/Na+ (pH2 = 10 Pa)
protein) (CooF), and a membrane-associated, 24–36 2x 2 H+
energy-converting [NiFe]-hydrogenase kDa [4Fe4S] Fd
ox
(CooHKLMUX), is coupled with chemios-
E' ≈ –500 mV
motic energy conservation as evidenced by
(Fd /Fd < 0.01)
ox red
growthanduncouplingexperiments(37–39).
Fd 2-
The energy-converting hydrogenases Cytoplasmic red
membrane
(EchA-F, EhaA-T, EhbA-Q, and MbhA-N)
+ –
org frommethanogenscontainsixconservedcore
ws. subunits (Figure 4) and up to 14 additional
e Figure4
viy. subunits.Thesixcoresubunitsshowsequence
ualree onl similarity to the six subunits of the carbon Scocnhveemratitnicgr[eNpirFesee]-nhtaytdiroongoefnathseessEtrcuhcAtu-rFe,EanhdaAfu-nTc,tEiohnboAf-Qth,eaennderMgyb-hA-N
nnus monoxide dehydrogenase (CooS)-associated foundinmethanogenicarchaea.Theenergy-convertinghydrogenaseEchA-F
w.anal [NiFe]-hydrogenase (CooHKLMUX) from iscomposedonlyofthesixconservedcoresubunits,whicharehighlightedin
wo
ed from w3. For pers bd(HaechytycedCrriDao,gEetFonGatsh)eef-raosfismvoecEia.sutceobldiu,[naNnitdisFteoo]f-shitxhyodefrtofhogeremncoaarsteee cafAuolbsnlobocrrtc.eioovTnniaht.ateTiionehnnse:eesFvregedsry,ua-fblceouhrrnnyedvidtersooraxtpirinhnegoswbyhiimtychdbartonowldgiozehen[yd4adFsbreeoys4paSEhr]hei-laacicsAluws-suTtibet,hruEsnd.hiatbssAhoe-fQdub,noaknundnodwManrbiehsA.-N
d1
wnloa06/10/ sduubcutansiets(NoufotAh-eNN)(AcoDmHp:luebxiIqoufinthoenreesopxiirdaotorrey- EchA-F. Genes for this type of energy-
Doon chain)fromE.coli(41).Oftheconservedsub- converting [NiFe]-hydrogenase are found in
7-536. Austin u(tnhiets,latrwgeor aorneeinmteogsrtalprmobeambblyraninevoplrvoetdeinins tMiv.orbaanrsk.eTrihaenydarMe.almsoazperiesbeunttninotthine gMen.oamcee-
79:50xas - cationtranslocation),andfourarehydrophilic of all members of the Methanomicrobiales.
10.Te proteins (Figure 4). Of the hydrophilic pro- The enzyme in these organisms is most
20of teins, one is the [NiFe]-hydrogenase large similar to the energy-converting hydrogenase
em. sity subunit, one the hydrogenase small subunit CooHKLMUX from R. rubrum (37) and
v. Biochy Univer woniteh),oonnlye oanne i[r4oFne-4suSl]f-ucrlusptreorte(tinhewpirtohxitmwaol Cho.whevyderr,ogeinnofonromtanfosrm(3in9g). aEcthigAh-tFcodmiffpelresx,
Reb [4Fe4S]-clusters, and one a subunit without withitsferredoxinandfunctionallyassociated
nu. a prosthetic group. In complex I, the subunit oxidoreductase.Thelackofcomplexformation
n
A NuoDhomologoustothe[NiFe]-hydrogenase is because in methanogens the reduced ferre-
large subunit lacks the N- and C-terminal doxin, generated by the energy-converting
CxxCmotifsfor[NiFe]-centerformation(35). hydrogenase, is used in electron transfer
None of the genes encoding energy- to more than one oxidoreductase. A 6 kDa
converting[NiFe]-hydrogenasesinBacteriaor 2[4Fe4S]-ferredoxin from M. barkeri, which
Archaea show a twin arginine translocation is most probably the ferredoxin reduced by
(Tat) motif-encoding sequence. The lack of H via the energy-converting hydrogenase
2
theTatmotif-encodingsequence(42)indicates EchA-F,hasbeencharacterized(43).
that the large subunit and the small subunit TheenzymecomplexEchA-Fhasbeenpu-
oftheenergy-converting[NiFe]-hydrogenases rified from M. barkeri and is composed of six
arenottranslocatedfromthecytoplasmtothe different subunits (Figure 4) encoded by the
Tat: twinarginine
periplasmandarethereforeorientedtowardthe echA-F operon (44, 45). EchA (69 kDa)
translocation
cytoplasm. and EchB (32 kDa) are the two integral
www.annualreviews.org • HydrogenasesfromMethanogens 515
ANRV413-BI79-18 ARI 27April2010 21:0
membraneproteins.EchAispredictedtohave site-directed mutants (54). Insights into the
17 membrane-spanning α-helices and shows mechanismofiontranslocationcomealsofrom
30% sequence identity to a putative Na+/H+ inhibition experiments with dicyclohexylcar-
translocatorcomponentinBacillussubtilis(44). bodiimide (DCCD), which specifically modi-
EchE is the large subunit, EchC is the small fies protonated carboxyl residues located in a
subunit,andtheseharborthe[NiFe]-centerand hydrophobicenvironment.Labelingstudiesof
a [4Fe4S]-cluster, respectively. EchF contains Echwith[14C]-DCCDshowedthattheinhibi-
two[4Fe4S]-clusters.EchDisthesolublesub- tionoftheenzymewasassociatedwithaspecific
unitwithoutaprostheticgroup.Chemicalanal- labeling of the two integral membrane sub-
ysesofthepurifiedcomplexhaverevealedthe units EchA and B, particularly of EchA (35).
presence of nickel, nonheme iron, and acid- TheinhibitionofEchbyDCCDindicatesthat
labile sulfur in a ratio of 1:12.5:12, substan- theelectrontransferreactioninthisenzymeis
tiating the presence of three [4Fe4S]-clusters strictlycoupledtocationtranslocation.
org in addition to the [NiFe]-center. Like CooH,
ws. thelargesubunitEchEissynthesizedwithouta EhaA-T and EhbA-Q. The [NiFe]-
e
viy. C-terminalextension;thegeneendsdirectlyaf- hydrogenase EhaA-T and EhbA-Q differ
ualree onl ter the DPCxxCxxR motif with a stop codon. from the energy-converting hydrogenase
nnus The evidence for reversed electron transfer EchA-Fbyhavingupto14additionalsubunits;
ww.aonal in ferredoxin reduction with H2 comes from manyofthesesubunitsareintegralmembrane
m wpers biochemical and genetic studies. Cell sus- proteins, and some are iron-sulfur proteins
wnloaded fro06/10/13. For pHmtieoi2ncns(CiEooOf(cid:4)onsCd=eOohf2y−dMt4ro1o.4gCbemOanraVks(ee)Er,,i(cid:4)ofiencr=varoteadll−voyinzx5ei2gn0,taahmnecdVytrE)oecpdwhluaiAtcsh--- (doa5lifsf5sofi)u.cbvuAualtnrfiiteutossnoecsnftiigcvooninsmiafigopcefal.entIxhtnlIeytoeafrwdetihdstteihitnoirogeunsltypa,ilartnahsuteoabrnpuyupncmaihtrsbeaneiinrst
Doon F. The reaction is driven by a proton motive changeinproperties;thus,complexIinE.coliis
7-536. Austin froeraccetio(4n6,)t.hTehdeehcyedllrsoaglesnoactiaotnalyozfeCtOhetroevCeOrse coofmmpooresetdhaonf4104ssuubbuunniittss(a5n6d,5in7)m. itochondria
10.79:50Texas - aanpdroHto2,nwmhoicthiveisfcoorucepl(e4d7,w4it8h).th(cid:2)eecbhumilduutpanot2sf areGfoeunnesdeinncoMd.inkganEdhlearAi,-Tinaanlld/moremEbhebrAs-Qof
20of did not catalyze the forward or the backward theMethanococcalesandMethanobacteriales,
em. sity reaction and also did not catalyze one of the and in some members of the Methanomicro-
v. Biochy Univer otitohneerdfaebrorevdeo(x4i9n,-5d0e)p.eTnhdeernetarreedcuocntifloinctsinmgerne-- Mbiaeltehsa(nbouretgnuolatibnooMneeit)h,aannodspehhaaearnudla/oparleuhstbrgiseanneds
Reb portswithrespecttothecouplingionusedby arenotfoundinmembersoftheMethanosarci-
nu. EchinM.barkeri.Theexperimentsinvestigat- nales.Insomemethanogens,e.g.,inM.marbur-
n
A ingthereversibleconversionofH andCO to gensis,thegenesareclusteredandformatran-
2 2
COandH Oaremoreinfavorofprotons(46– scriptionunit,andinothers,e.g.,M.jannaschii,
2
48),whereasthoseaddressingthereductionof someofthegenesareinseparateloci.
CO withH toformylmethanofuranaremore Theehaoperon(12.5kb)inM.marburgensis
2 2
infavorofsodiumions(51,52). is composed of 20 open reading frames that
The iron-sulfur centers in the EchA-F form a transcription unit (55). Sequence
complex have been characterized by electron analysisindicatesthatfourofthegenesencode
paramagnetic resonance spectroscopy, reveal- proteinswithhighsequencesimilaritytofour
ing that two of the [4Fe4S]-clusters show of the six different subunits characteristic for
pH-dependent redox potentials (53) and in- energy-converting hydrogenases (Figure 4):
dicating that these clusters mediate elec- ehaO encodes the large [NiFe]-center harbor-
tronandprotontransfer.The[4Fe4S]-clusters ingsubunitasapreprotein;ehaNencodesthe
were assigned to the individual subunits via small subunit with only one [4Fe4S]-cluster.
516 Thaueretal.