α-D-galactosidaseactivityandgalactomannanandgalactosylsucroseoligosaccharidedepletioningerminatinglegumeseeds.
McCleary,B.V.&Matheson,N.K.(1974).Phytochemistry,13(9),1747-1757.
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Germinatingseedsoflucerne,guar,carobandsoybeaninitiallydepletedraffinoseseriesoligosaccharidesandthengalactomannan.Thisdepletionwasaccompaniedbyarapidincreaseandthenadecreaseinα-galactosidaselevels.Lucerneandguarcontainedtwoα-galactosidaseactivities,carobthreeandsoybeanfour.Oneoftheseineachplant,fromitslocationintheendosperm,timeofappearanceandkineticbehaviour,appearedtobeprimarilyinvolvedingalactomannanhydrolysis.ThisenzymeinlucernehadMWof23000andcouldnotbeseparatedfromβ-mannanaseby(NH4)2SO4fractionation,DEAE,CMorSE-cellulosechromatographyorgelfiltration,butonlybypolyacrylamidegelelectrophoresis.Inguar,carobandsoybean,itcouldbeseparatedbyion-exchangechromatographyandgelfiltration.Inlucerne,carobandguarmostofthetotalincreaseinactivitywasduetothisenzyme.Theotherα-galactosidaseshadMWsofabout35000andcouldbeseparatedfromβ-mannanasebydissection,ionexchangecellulosechromatographyandgelfiltration.Theywerelocatedinthecotyledon-embryoandappearedtobeprimarilyinvolvedingalactosylsucroseoligosaccharidehydrolysis.
Galactomannanstructureandβ-mannanaseandβ-mannosidaseactivityingerminatinglegumeseeds.
McCleary,B.V.&Matheson,N.K.(1975).Phytochemistry,14(5-6),1187-1194.
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Structuralchangesingalactomannanongerminationoflucerne,carob,honeylocust,guarandsoybeanseeds,asmeasuredbyviscosity,elutionvolumesongelfiltrationandultra-centrifugationwereslightconsistentwitharapidandcompletehydrolysisofamoleculeoncehydrolysisofthemannanchainstarts.β-Mannanaseactivityincreasedandthendecreased,parallelinggalactomannandepletion.Multipleformsofβ-mannanasewereisolatedandthesewerelocatedintheendosperm.β-Mannanasehadlimitedabilitytohydrolysegalactomannanswithhighgalactosecontents.Seedscontainingthesegalactomannanshadveryactiveα-galactosidases.β-Mannosidaseswerepresentinbothendospermandcotyledon-embryoandcouldbeseparatedchromatographically.Thelevelofactivitywasjustsufficienttoaccountformannoseproductionfrommanno-oligosaccharides.
Galactomannansandagalactoglucomannaninlegumeseedendosperms:Structuralrequirementsforβ-mannanasehydrolysis.
McCleary,B.V.,Matheson,N.K.&Small,D.B.(1976).Phytochemistry,15(7),1111-1117.
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Aseriesofgalactomannanswithvaryingdegreesofgalactosesubstitutionhavebeenextractedfromtheendospermsoflegumeseedswithwaterandalkaliandtheamountofsubstitutionrequiredforwatersolubilityhasbeendetermined.Somewereheterogeneouswithrespecttothedegreeofgalactosesubstitution.Thestructuralrequirementsforhydrolysisbyplantβ-mannanasehavebeenstudiedusingtherelativeratesandextentsofhydrolysisofthesegalactomannans.Amoredetailedexaminationoftheproductsofhydrolysisofcarobgalactomannanhasbeenmade.Atleasttwocontiguousanhydromannoseunitsappeartobeneededforscission.Thisissimilartotherequirementforhydrolysisbymicrobialenzymes.Judastree(Cercissiliquastrum)endospermcontainedapolysaccharidewithauniquecompositionforalegumeseedreserve.Gelchromatographyandelectrophoresisoncelluloseacetateindicatedhomogeneity.Hydrolysiswithamixtureofβ-mannanaseandα-galactosidasegaveaglucose-mannosedisaccharideandacetolysisgaveagalactose-mannose.Theseresults,aswellasthepatternofhydrolysisbyβ-mannanasewereconsistentwithagalactoglucomannanstructure.
Modesofactionofβ-mannanaseenzymesofdiverseoriginonlegumeseedgalactomannans.
McCleary,B.V.(1979).Phytochemistry,18(5),757-763.
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β-MannanaseactivitiesinthecommercialenzymepreparationsDriselaseandCellulase,inculturesolutionsofBacillussubtilis(TX1),incommercialsnailgut(Helixpomatia)preparationsandingerminatedseedsoflucerne,Leucaenaleucocephalaandhoneylocust,havebeenpurifiedbysubstrateaffinitychromatographyonglucomannan-AH-Sepharose.Onisoelectricfocusing,multipleproteinbandswerefound,allofwhichhadβ-mannanaseactivity.EachpreparationappearedasasinglemajorbandonSDS-polyacrylamidegelelectrophoresis.Theenzymesvariedintheirfinalspecificactivities,Kmvalues,optimalpH,isoelectricpointsandpHandtemperaturestabilitiesbuthadsimilarMWs.Theenzymeshavedifferentabilitiestohydrolysegalactomannanswhicharehighlysubstitutedwithgalactose.ThepreparationsDriselaseandCellulasecontainβ-mannanaseswhichcanattackhighlysubstitutedgalactomannansatpointsofsingleunsubstitutedD-mannosylresiduesiftheD-galactoseresiduesinthevicinityofthebondtobehydrolysedareallononlyonesideofthemainchain.
AnenzymictechniqueforthequantitationofgalactomannaninguarSeeds.
McCleary,B.V.(1981).Lebensmittel-Wissenschaft&Technologie,14,56-59.
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Anenzymictechniquehasbeendevelopedfortherapidandaccuratequantitationofthegalactomannancontentofguarseedsandmillingfractions.Thetechniqueinvolvesthemeasurementofthegalactosecomponentofgalactomannansusinggalactosedehydrogenase.Thegalactomannansareconvertedtogalactoseandmanno-oligosaccharidesusingpartiallypurifiedenzymesfromacommercialpreparationandfromgerminatedguarseeds.Simpleprocedureshavebeendevisedforthepreparationoftheseenzymes.Applicationofthetechniquetoanumberofguarvarietiesgavevaluesforthegalactomannancontentrangingfrom22.7to30.8%ofseedweight.
Purificationandpropertiesofaβ-D-mannosidemannohydrolasefromguar.
McCleary,B.V.(1982),CarbohydrateResearch,101(1),75-92.
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Aβ-D-mannosidemannohydrolaseenzymehasbeenpurifiedtohomogeneityfromgerminatedguar-seeds.Difficultiesassociatedwiththeextractionandpurificationappearedtobeduetoaninteractionoftheenzymewithotherproteinmaterial.Thepurifiedenzymehydrolysedvariousnaturalandsyntheticsubstrates,includingβ-D-manno-oligosaccharidesandreducedβ-D-manno-oligosaccharidesofdegreeofpolymerisation2to6,aswellasp-nitrophenyl,naphthyl,andmethylumbelliferylβ-D-mannopyranosides.Thepreferred,naturalsubstratewasβ-D-mannopentaose,whichwashydrolysedattwicetherateofβ-D-mannotetraoseandfivetimestherateofβ-D-mannotriose.Thisresult,togetherwiththeobservationthatα-D-mannoseisreleasedonhydrolysis,indicatesthattheenzymeisanexo-β-D-mannanase.
Preparative–scaleisolationandcharacterisationof61-α-D-galactosyl-(1→4)-β-D-mannobioseand62-α-D-galactosyl-(1→4)-β-D-mannobiose.
McCleary,B.V.,Taravel,F.R.&Cheetham,N.W.H.(1982).CarbohydrateResearch,104(2),285-297.
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N.m.r.,enzymic,andchemicaltechniqueshavebeenusedtocharacterisetheD-galactose-containingtri-andtetra-saccharidesproducedonhydrolysisofcaroband
L.leucocephalaD-galacto-D-mannansbyDriselaseβ-D-mannanase.Theseoligosaccharideswereshowntobeexclusively6
1-α-D-galactosyl-β-D-mannobioseand6
1-α-D-galactosyl-β-D-mannotriose.Fur
Thermore,theseweretheonlyD-galactose-containingtri-andtetra-saccharidesproducedonhydrolysisofcarobD-galacto-D-mannanbyβ-D-mannanasesfromothersources,including
Bacillussubtilis,
Aspergillusniger,
Helixpomatiagutsolution,andgerminatedlegumes.Acidhydrolysisoflucernegalactomannanyielded6
1-α-D-galactosyl-β-D-mannobioseand6
2-α-D-galactosyl-β-D-mannobiose.
β-D-mannosidasefromHelixpomatia.
McCleary,B.V.(1983).CarbohydrateResearch,111(2),297-310.
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β-D-Mannosidase(β-D-mannosidemannohydrolaseEC3.2.1.25)waspurified160-foldfromcrudegut-solutionof
Helixpomatiabythreechromatographicstepsandthengaveasingleproteinband(mol.wt.94,000)onSDS-gelelectrophoresis,andthreeproteinbands(ofalmostidenticalisoelectricpoints)onthin-layeriso-electricfocusing.Eachoftheseproteinbandshadenzymeactivity.Thespecificactivityofthepurifiedenzymeon
p-nitrophenylβ-D-mannopyranosidewas1694nkat/mgat40°anditwasdevoidofα-D-mannosidase,β-D-galactosidase,2-acet-amido-2-deoxy-D-glucosidase,(1→4)-β-D-mannanase,and(1→4)-β-D-glucanaseactivities,almostdevoidofα-D-galactosidaseactivity,andcontaminatedwith<0.02% of="" β-d-glucosidase="" activity.="" the="" purified="" enzyme="" had="" the="" same="">0.02%>
Kmforborohydride-reducedβ-D-manno-oligosaccharidesofd.p.3-5(12.5mM).Theinitialrateofhydrolysisof(1→4)-linkedβ-D-manno-oligosaccharidesofd.p.2-5andofreducedβ-D-manno-oligosaccharidesofd.p.3-5wasthesame,and
o-nitrophenyl,methylumbelliferyl,andnaphthylβ-D-mannopyranosideswerere
ADIlyhydrolysed.β-D-Mannobiosewashydrolysedatarate~25timesthatof6
1-α-D-galactosyl-β-D-mannobioseand6
3-α-D-galactosyl-β-D-mannotetraose,andat~90timestherateforβ-D-mannobi-itol.
Enzymicinteractionsinthehydrolysisofgalactomannaningerminatingguar:Theroleofexo-β-mannanase.
McCleary,B.V.(1983).Phytochemistry,22(3),649-658.
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Hydrolysisofgalactomannaninendospermsofgerminatingguarisduetothecombinedactionofthreeenzymes,α-galactosidase,β-mannanaseandexo-β-mannanase.α-Galactosidaseandexo-β-mannanaseactivitiesoccurbothinendospermandcotyledontissuebutβ-mannanaseoccursonlyinendosperms.Onseedgermination,β-mannanaseandendospermicα-galactosidasearesynthesizedandactivitychangesparallelgalactomannandegradation.Galactomannandegradationandsynthesisofthesetwoenzymesareinhibitedbycycloheximide.Incontrast,endospermicexo-β-mannanaseisnotsynthesizedonseedgermination,butratherisalreadypresentthroughoutendospermtissue.Ithasnoactiononnativegalactomannan.α-Galactosidase,β-mannanaseandexo-β-mannanasehavebeenpurifiedtohomogeneityandtheirseparateandcombinedactioninthehydrolysisofgalactomannanandeffectontherateofuptakeofcarbohydratebycotyledons,studied.Resultsobtainedindicatedthatthesethreeactivitiesaresufficienttoaccountforgalactomannandegradationinvivoand,further,thatallthreearerequired.Cotyledonscontainanactiveexo-β-mannanaseandsugar-uptakeexperimentshaveshownthatcotyledonscanabsorbmannobioseintact,indicatingthatthisenzymeisinvolvedinthecompletedegradationofgalactomannanonseedgermination.
Characterisationoftheoligosaccharidesproducedonhydrolysisofgalactomannanwithβ-D-mannase.
McCleary,B.V.,Nurthen,E.,Taravel,F.R.&Joseleau,J.P.(1983).CarbohydrateResearch,118,91-109.
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Treatmentofhot-water-solublecarobgalactomannanwithβ-D-mannanasesfromA.nigerorlucerneseedaffordsanarrayofD-galactose-containingβ-D-mannosaccharidesaswellasβ-D-manno-biose,-triose,and-tetraose(lucerne-seedenzymeonly).TheD-galactose-containingβ-D-mannosaccharidesofd.p.3–9producedbyA.nigerβ-D-mannanasehavebeencharacterised,usingenzymic,n.m.r.,andchemicaltechniques,as61-α-D-galactosyl-β-D-mannobiose,61-α-D-galactosyl-β-D-mannotriose,63,64-di-α-D-galactosyl-β-D-mannopentaose(theonlyheptasaccharide),and63,64-di-α-D-galactosyl-β-D-mannohexaose,64,65-di-α-D-galactosyl-β-D-mannohexaose,and61,63,64-tri-α-D-galactosyl-β-D-mannopentaose(theonlyoctasaccharides).Fournonasaccharideshavealsobeencharacterised.Penta-andhexa-saccharideswereabsent.Lucerne-seedβ-D-mannanaseproducedthesamebranchedtri-,tetra-andhepta-saccharides,andalsopenta-andhexa-saccharidesthatwerecharacterisedas61-α-D-galactosyl-β-D-mannotetraose,63-α-D-galactosyl-β-D-mannotetraose,61,63-di-α-D-galactosyl-β-D-mannotetraose,63-α-D-galactosyl-β-D-mannopentaose,and64-α-D-galactosyl-β-D-mannopentaose.NoneoftheoligosaccharidescontainedaD-galactosestubontheterminalD-mannosylgroupnorweretheysubstitutedonthesecondD-mannosylresiduefromthereducingterminal.
Actionpatternsandsubstrate-bindingrequirementsofβ-D-mannanasewithmannosaccharidesandmannan-typepolysaccharides.
McCleary,B.V.&Matheson,N.K.(1983).CarbohydrateResearch,119,191-219.
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Purified(1→4)-β-D-mannanasefromAspergillusnigerandlucerneseedshasbeenincubatedwithmannosaccharidesandend-reduced(1→4)-β-D-mannosaccharidesand,fromtheproductsofhydrolysis,acyclicreaction-sequencehasbeenproposed.Fromtheheterosaccharidesreleasedbyhydrolysisofthehot-water-solublefractionofcarobgalactomannanbyA.nigerβ-D-mannanase,apatternofbindingbetweentheβ-D-mannanchainandtheenzymehasbeendeduced.Theproductsofhydrolysiswiththeβ-D-mannanasesfromIrpexlacteus,Helixpomatia,Bacillussubtilis,andlucerneandguarseedshavealsobeendetermined,andthedifferencesfromtheactionofA.nigerβ-D-mannanaserelatedtominordifferencesinsubstratebinding.Theproductsofhydrolysisofglucomannanareconsistentwiththoseexpectedfromthebindingpatternproposedfromthehydrolysisofgalactomannan.
Thefinestructuresofcarobandguargalactomannans.
McCleary,B.V.,Clark,A.H.,Dea,I.C.M.&Rees,D.A.(1985).CarbohydrateResearch,139,237-260.
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ThedistributionofD-galactosylgroupsalongtheD-mannanbackbone(finestructure)ofcarobandguargalactomannanshasbeenstudiedbyacomputeranalysisoftheamountsandstructuresofoligosaccharidesreleasedonhydrolysisofthepolymerswithtwohighlypurifiedβ-D-mannanasesisolatedfromgerminatedguarseedandfromAspergillusnigercultures.Computerprogrammesweredevelopedwhichaccountedforthespecificsubsite-bindingrequirementsoftheβ-D-mannanasesandwhichsimulatedthesynthesisofgalactomannanbyprocessesinwhichtheD-galactosylgroupsweretransferredtothegrowingD-mannanchainineitherastatisticallyrandommannerorasinfluencedbynearest-neighbour/second-nearest-neighboursubstitution.Suchamodelwaschosenasitisconsistentwiththeknownpatternofsynthesisofsimilarpolysaccharides,forexample,xyloglucan;also,additiontoapreformedmannanchainwouldbeunlikely,duetotheinsolublenatureofsuchpolymers.TheD-galactosedistributionincarobgalactomannanandinthehot-andcold-water-solublefractionsofcarobgalactomannanhasbeenshowntobenon-regular,withahighproportionofsubstitutedcouplets,lesseramountsoftriplets,andanabsenceofblocksofsubstitution.TheprobabilityofsequencesinwhichalternateD-mannosylresiduesaresubstitutedislow.TheprobabilitydistributionofblocksizesforunsubstitutedD-mannosylresiduesindicatesthatthereisahigherproportionofblocksofintermediatesizethanwouldbepresentinagalactomannanwithastatisticallyrandomD-galactosedistribution.Basedonthealmostidenticalpatternsofamountsofoligosaccharidesproducedonhydrolysiswithβ-D-mannanase,itappearsthatgalactomannansfromseedofawiderangeofcarobvaritieshavethesamefine-structure.TheD-galactosedistributioninguar-seedgalactomannanalsoappearstobenon-regular,andgalactomannansfromdifferentguar-seedvarietiesappeartohavethesamefine-structure.
Effectofgalactose-substitution-patternsontheinteractionpropertiesofgalactomannas.
Dea,I.C.M.,Clark,A.H.&McCleary,B.V.(1986).CarbohydrateResearch,147(2),275-294.
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ArangeofgalactomannansvaryingwidelyinthecontentsofD-galactosehavebeencomparedforself-associationandtheirinteractionpropertieswithagaroseandxanthan.Whereas,ingeneral,themostinteractivegalactomannansarethoseinwhichthe(1→4)-β-D-mannanchainisleastsubstitutedbyα-D-galactosylstubs,evidenceispresentedwhichindicatesthatthedistributionofD-galactosylgroupsalongthebackbone(finestructure)canhaveasignificanteffectontheinteractionproperties.Forgalactomannanscontaining<30% of="" d-galactose,="" those="" which="" contain="" a="" higher="" frequency="" of="" unsubstituted="" blocks="" of="" intermediate="" length="" in="" the="" β-d-mannan="" chain="" are="" most="" interactive.="" for="" galactomannans="" containing="">40%ofD-galactose,thosewhichcontainahigherfrequencyofexactlyalternatingregionsintheβ-D-mannanchainaremostinteractive.Thisselectivity,onthebasisofgalactomannanfine-structure,inmixedpolysaccharideinteractions
invitrocouldmimictheselectivityofbindingofbranchedplant-cell-wallpolysaccharidesin
BIOLOGicalsystems.30%>
Effectofthemolecularfinestructureofgalactomannansontheirinteractionproperties-theroleofunsubstitutedsides.
Dea,I.C.M.,Clark,A.H.&McCleary,B.V.(1986).FoodHydrocolloids,1(2),129-140.
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ArangeofgalactomannansvaryingwidelyinthecontentofD-galactosehavebeencomparedforself-association,andtheirinteractionpropertieswithagaroseandxanthan.TheresultspresentedindicatethatingeneralthemostinteractivegalactomannansarethoseinwhichtheD-mannanmainchainbearsfewestD-galactosestubs,andconfirmthatthedistributionofD-galactosegroupsalongthemainchaincanhaveasignificanteffectontheinteractivepropertiesofthegalactomannans.Ithasbeenshownthatfreeze—thawprecipitationofgalactomannansrequiresregionsoftotallyunsubstitutedD-mannoseresiduesalongthemainchain,andthatathresholdforsignificantfreeze—thawprecipitationoccursataweight-averagelengthoftotallyunsubstitutedresiduesofapproximatelysix.ForgalactomannanshavingstructuresabovethisthresholdtheirinteractivepropertieswithotherpolysaccharidesarecontrolledbystructuralfeaturesassociatedwithtotallyunsubstitutedregionsoftheD-mannanbackbone.Incontrast,forgalactomannansbelowthisthreshold,theirinteractivepropertiesarecontrolledbystructuralfeaturesassociatedwithunsubstitutedsidesofD-mannanbackbone.
GalactomannanchangesindevelopingGleditsiaTriacanthosSeeds.
McCleary,B.V.,Mallett,I.&Matheson,N.K.(1987).Phytochemistry,26(7),1889-1894.
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Galactomannanhasbeenextractedfromtheendospermofseedsof
Gleditsiatriacanthos(honeylocust)atdifferentstagesofdevelopment,whentheseedwasaccumulatingstoragematerial.Propertiesofthedifferentsampleshavebeenstudied.Themolecularsizedistributionbecamemoredisperseasgalactomannanaccumulatedandthegalactose:mannoseratiodecreasedslightly.Someposs
IBLereasonsforthesechangesarediscussed.
Distributionandcharacterisationoffructaninwheatmillingfractions.
Haskå,L.,Nyman,M.&Andersson,R.(2008).JournalofCerealScience,48(3),768-774.
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Structureandhealtheffectsofinulin-typefructanshavebeenextensivelystudied,whilelessisknownaboutthepropertiesofthegraminan-typefructansinwheat.Arabinoxylan(AX)isanotherimportantindigestiblecomponentincerealgrains,whichmayhavebeneficialhealtheffects.Inthisstudy,thefructancontentinmillingfractionsoftwowheatcultivarswasdeterminedandrelatedtoash,dietaryfibreandAXcontents.ThemolecularweightdistributionofthefructanswasanalysedwithHPAEC-PADandMALDI-TOFMSusing1HNMRandenzymatichydrolysisforidentificationoffructans.Thefructancontent(g/100g)rangedfrom1.5±0.2inflourto3.6±0.5inshortsand3.7±0.3inbran.Acorrelationwasfoundbetweenfructancontentanddietaryfibrecontent(r=0.93,P<0.001),=""but=""with=""a=""smaller=""variation=""in=""fructan=""content=""between=""inner=""and=""outer=""parts=""of=""the=""grain.=""about=""50%=""of=""the=""dietary=""fibre=""consisted=""of=""ax=""in=""all=""fractions.=""the=""fructans=""were=""found=""to=""have=""a=""dp=""of=""up=""to=""19=""with=""a=""similar=""molecular=""weight=""distribution=""in=""the=""different=""fractions.="">
Effectofabandonmentandplantclassificationoncarbohydratereservesofmeadowplants.
Janeček,Š.,Lanta,V.,Klimešová,J.&Doležal,J.(2011).PlantBiology,13(2),243-251.
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Westudiedtheeffectofcessationofmanagementoncarbohydratereservesofplantsinmeadowswithdifferentenvironmentalcharacteristicsandplantcomposition.Werecordedstoragecarbohydratesandseasonalchangesfor40plantspecies.Weaskedwhethertherearedifferencesinresponsesofcarbohydratereservesinforbsversusgraminoidsandinplantsstoringstarchversusplantsstoringosmoticallyactivecarbohydrates.Weanalysedbelowgroundorgansbeforethemeadowsweremownandattheendofthevegetationseasoninmownversusrecentlyabandonedplots.Whereasstarchandfructanswerewidelydistributed,raffinosefamilyoligosaccharideswerethemaincarbohydratereservesoftheLamiaceaeandPlantagolanceolata.Propertiesofcarbohydratereservesdifferedbetweenforbsandgraminoidsbutnodifferencewasfoundbetweenplantsstoringstarchversusosmoticallyactivecarbohydrates.Graminoidshadlowercarbohydrateconcentrationsthanforbs.Weobservedapositiveeffectofmowingoncarbohydrateconcentrationsofgraminoidsinthedry,calcium-richmeadowandhigherseasonalfluctuationsofthesevaluesintheacid,wetmeadow,suggestingthatlocalfactorsand/orthespeciespoolaffectcarbohydratereserves.Despitelocalconditions,graminoidsrepresentadistinctfunctionalgroupinmeadowsfromthepointofviewoftheirstorageeconomy.Wesuggestthataswellasgrowth,storageprocessesshouldalsobeconsideredforunderstandingthefunctioningofmeadowplantcommunities.