TheL-ArABInose/D-Galactosetestkitisa simple,reliableandaccurateUVmethodforthemeasurementandanalysisofL-arabinoseand/orD-galactoseinvariousmaterialsincludingfoods,feeds,beveragesandplantproducts. Extendedcofactorsstability.Dissolvedcofactorsstablefor>1yearat4oC. Suitableformanual,auto-analyserandmicroplateformats.Hydrolysisofwheatflourarabinoxylan,acid-debranchedwheatflourarabinoxylanandarabino-xylo-oligosaccharidesbyβ-xylanase,α-L-arabinofuranosidaseandβ-xylosidase.McCleary,B.V.,McKie,V.A.,Draga,A.,Rooney,E.,Mangan,D.&Larkin,J.(2015).CarbohydrateResearch,407,79-96.LinktoArticleReadAbstractArangeofα-L-arabinofuranosyl-(1-4)-β-D-xylo-oligosaccharides(AXOS)wereproducedbyhydrolysisofwheatflourarabinoxylan(WAX)andaciddebranchedarabinoxylan(ADWAX),inthepresenceandabsenceofanAXH-d3α-L-arabinofuranosidase,byseveralGH10andGH11β-xylanases.ThestructuresoftheoligosaccharideswerecharacterisedbyGC-MSandNMRandbyhydrolysisbyarangeofα-L-arabinofuranosidasesandβ-xylosidase.TheAXOSwerepurifiedandusedtocharacterisetheactionpatternsofthespecificα-L-arabinofuranosidases.Theseenzymes,incombinationwitheitherCellvibriomixtusorNeocallimastixpatriciarumβ-xylanase,wereusedtoproduceelevatedlevelsofspecificAXOSonhydrolysisofWAX,suchas32-α-L-Araf-(1-4)-β-D-xylobiose(A3X),23-α-L-Araf-(1-4)-β-D-xylotriose(A2XX),33-α-L-Araf-(1-4)-β-D-xylotriose(A3XX),22-α-L-Araf-(1-4)-β-D-xylotriose(XA2X),32-α-L-Araf(1-4)-β-D-xylotriose(XA3X),23-α-L-Araf-(1-4)-β-D-xylotetraose(XA2XX),33-α-L-Araf-(1-4)-β-D-xylotetraose(XA3XX),23,33-di-α-L-Araf-(1-4)-β-D-xylotriose(A2+3XX),23,33-di-α-L-Araf-(1-4)-β-D-xylotetraose(XA2+3XX),24,34-di-α-L-Araf-(1-4)-β-D-xylopentaose(XA2+3XXX)and33,34-di-α-L-Araf-(1-4)-β-D-xylopentaose(XA3A3XX),manyofwhichhavenotpreviouslybeenproducedinsufficientquantitiestoallowtheiruseassubstratesinfurtherenzymicstudies.ForA2,3XX,yieldsofapproximately16%ofthestartingmaterial(wheatarabinoxylan)havebeenachieved.Mixturesoftheα-L-arabinofuranosidases,withspecificactiononAXOS,havebeencombinedwithβ-xylosidaseandβ-xylanasetoobtainanoptimalmixtureforhydrolysisofarabinoxylantoL-arabinoseandD-xylose.Novelbifunctionalα-L-arabinofuranosidase/xylobiohydrolase(ABF3)fromPenicilliumpurpurogenum.Ravanal,M.C.,Callegari,E.&Eyzaguirre,J.(2010).AppliedandEnvironmentalMicroBIOLOGy,76(15),5247-5253.LinktoArticleReadAbstractThesoftrotfungusPenicilliumpurpurogenumgrowsonavarietyofnaturalsubstratesandsecretesvariousisoformsofxylanolyticenzymes,includingthreearabinofuranosidases.Thisworkdescribesthebiochemicalpropertiesaswellasthenucleotideandaminoacidsequencesofarabinofuranosidase3(ABF3).Thisenzymehasbeenpurifiedtohomogeneity.Itisaglycosylatedmonomerwithamolecularweightof50,700andcanbindcellulose.Theenzymeisactivewithp-nitrophenylα-L-arabinofuranosideandp-nitrophenylβ-D-xylopyranosidewithAKmof0.65mMand12mM,respectively.Theenzymeisactiveonxylooligosaccharides,yieldingproductsofshorterlength,includingxylose.However,itdoesnothydrolyzearabinooligosaccharides.Whenassayedwithpolymericsubstrates,littlearabinoseisliberatedfromarabinananddebranchedarabinan;however,ithydrolyzesarabinoseandreleasesxylooligosaccharidesfromarabinoxylan.SequencingbothABF3CDNAandgenomicDNArevealsthatthisgenedoesnotcontainintronsandthattheopenreADIngframeis1,380nucleotidesinlength.Thededucedmatureproteiniscomposedof433aminoacidsresiduesandhasacalculatedmolecularweightof47,305.ThededucedaminoacidsequencehasbeenvalidatedbymassspectrometryanalysisofpeptidesfrompurifiedABF3.Atotalof482bpofthepromoterweresequenced;putativebindingsitesfortranscriptionfactorssuchasCreA(four),XlnR(one),andAreA(three)andtwoCCAATboxeswerefound.Theenzymehastwodomains,onesimilartoproteinsofglycosylhydrolasefamily43attheamino-terminalendandafamily6carbohydratebindingmoduleatthecarboxylend.ABF3isthefirstdescribedmodularfamily43enzymefromafungalsource,havingbothα-L-arabinofuranosidaseandxylobiohydrolasefunctionalities.TheinfluenceofAspergillusnigertranscriptionfactorsAraRandXlnRinthegeneexpressionduringgrowthinD-xylose,L-arabinoseandsteam-explodedsugarcanebagasse.deSouza,W.R.,Maitan-Alfenas,G.P.,deGouvêa,P.F.,Brown,N.A.,Savoldi,M.,Battaglia,E.,GoldmanM.H.S.,deVries,R.P.&Goldman,G.H.(2013).FungalGeneticsandBiology,60,29-45.LinktoArticleReadAbstractTheinterestintheconversionofplantbiomasstorenewablefuelssuchasbioethanolhasledtoanincreasedinvestigationintotheprocessesregulatingbiomasssaccharification.ThefilamentousfungusAspergillusnigerisanimportantmicroorganismcapableofproducingawidevarietyofplantbiomassdegradingenzymes.InA.nigerthetranscriptionalactivatorXlnRanditsclosehomolog,AraR,controlsthemain(hemi-)cellulolyticsystemresponsIBLeforplantpolysaccharidedegradation.Sugarcaneisusedworldwideasafeedstockforsugarandethanolproduction,whilethelignocellulosicresidualbagassecanbeusedindifferentindustrialapplications,includingethanolproduction.Theuseofpentosesugarsfromhemicellulosesrepresentsanopportunitytofurtherincreaseproductionefficiencies.Inthepresentstudy,wedescribeaglobalgeneexpressionanalysisofA.nigerXlnR-andAraR-deficientmutantstrains,grownonaD-xylose/L-arabinosemonosaccharidemixtureandsteam-explodedsugarcanebagasse.DifferentgenesetsofCAZyenzymesandsugartransporterswereshowntobeindividuallyorduallyregulatedbyXlnRandAraR,withXlnRappearingtobethemajorregulatoroncomplexpolysaccharides.Ourstudycontributestounderstandingofthecomplexregulatorymechanismsresponsibleforplantpolysaccharide-degradinggeneexpression,andopensnewpossibilitiesfortheengineeringoffungiabletoproducemoreefficientenzymaticcocktailstobeusedinbiofuelproduction.HomologousexpressionandbiochemicalcharacterizationofthearylsulfatasefromKluyveromyceslactisanditsrelevanceinmilkprocessing.Stressler,T.,Leisibach,D.,Lutz-Wahl,S.,Kuhn,A.&Fischer,L.(2016).AppliedMicrobiologyandBiotechnology,100(12),5401-5414.LinktoArticleReadAbstractTheindustrialmanufacturingprocessoflactose-freemilkproductsdependsontheapplicationofcommercialβ-galactosidase(lactase)preparations.ThesepreparationsareoftenobtainedfromKluyveromyceslactis.ThereisagenepresentinthegenomeofK.lactiswhichshouldencodeforanenzymecalledarylsulfatase(EC3.1.6.1).Therefore,thisenzymecouldalsobepresentinβ-galactosidasepreparations.ThearylsulfataseissUSPectedofbeingresponsibleforanunpleasant“cowshed-like”off-flavorresultingfromthereleaseofp-cresolfrommilkendogenousalkylphenolsulfuricesters.Sofar,nogene/functionalityrelationshipisdescribed.Inaddition,nostudyisavailablewhichhasshownthatarylsulfatasefromK.lactisistrulyresponsiblefortheflavorgeneration.Inthisstudy,weclonedtheputativearylsulfatasegenefromK.lactisGG799intothecommerciallyavailablevectorpKLAC2.Thecloningstrategychosenresultedinahomologous,secretoryexpressionofthearylsulfatase.Weshowedthattheheretoforeputativearylsulfatasehasthedesiredactivitywiththesyntheticsubstratep-nitrophenylsulfateandwiththenaturalsubstratep-cresolsulfate.Theenzymewasbiochemicallycharacterizedandshowedanoptimumtemperatureof45-50 CandanoptimumpHof9-10.Additionally,thearylsulfatasewasactivatedbyCa2+ionsandwasinactivatedbyZn2+ions.Moreover,thearylsulfatasewasinhibitedbyp-cresolandsulfateions.Finally,theenzymewasaddedtoultra-heattreated(UHT)milkandasensorytriangletestverifiedthatthearylsulfatasefromK.lactiscancauseanunpleasant“cowshed-like”off-flavor.UV-methodforthedeterminationofL-ArabinoseandD-GalactoseinhydrolysedplantproductsPrinciple:                   (galactosemutarotase)(1)α-L-Arabinose/α-D-galactose↔β-L-arabinose/β-D-galactose           (β-galactosedehydrogenase)(2)β-L-Arabinose+NAD+→L-arabinonicacid+NADH+H+                     (β-galactosedehydrogenase)(3)β-D-Galactose+NAD+→D-galactonicacid+NADH+H+Kitsize:                         *115assays(manual)/1150(microplate)                                        /1150(auto-analyser)* Thenumberofmanualtestsperkitcanbedoubledifallvolumesarehalved. ThiscanbereadilyaccommodatedusingtheMegaQuantTM WaveSpectrophotometer(D-MQWAVE).Method:                          Spectrophotometricat340nmReactiontime:                 ~12min(L-arabinose)~6min(D-galactose)Detectionlimit:                0.58mg/L(L-arabinose)0.69mg/L(D-galactose)Applicationexamples:Analysisofhydrolysatesofoligo-andpolysaccharides(e.g.arabinan,arabinoxylan,galactan,arabinogalactan),milk,dairyproducts,foodscontainingmilk(e.g.dieteticfoods,bakeryproducts,babyfood,chocolate,sweetsandice-cream),foodadditives(e.g.sweeteners),cosmetics,pharmaceuticalsandothermaterials(e.g.biologicalcultures,samples,etc.)Methodrecognition:     NovelmethodAdvantagesVeryrapidreactionduetoinclusionofgalactosemutarotase(patentedtechnology) Verycosteffective Allreagentsstablefor>2yearsafterpreparation Onlyenzymatickitavailable Mega-Calc™softwaretoolisavailablefromourwebsiteforhassle-freerawdataprocessing Standardincluded Suitableformanual,microplateandauto-analyserformats

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