Description

Theenzymereleasesnon-reducingterminalα(1-3,4,6)-linkedgalactosefromoligosaccharides.

Source:
Greencoffeebean.

NOTE:Highlypurified;β-galactosidasecontaminatingactivityisnotdetectable,asdeterminedbyextendedincubationwith2-ABNA2(G2).

Specificity:
Coffeebeanα-galactosidasecleavesnon-reducingterminalgalactoseresidueslinkedα1-3,α1-4andα1-6topolysaccharides,glycoproteinsandglycans.Theprecisespecificityoftheenzymeissomewhatdependentonthenatureoftheglycoconjugate.

Figure1:CleavagespecificityforGKX-5007 α(1-3,4,6)-Galactosidase.

Applications:
Terminalα-linkedgalactoseresidueshavebeenreportedtooccuronseveralglycoproteinsandareofincreasinginterest.Thegalactose-α-1-3Galgroupiswidelyfoundinglycoconjugatesfromnon-primatemammalsandNewWorldmonkeysalthoughitisabsentfromhumansandOldWorldmonkeys.Thepotentialpresenceofthismodificationanditsassociatedantigenicityisanimportantconsiderationintheexpressionofrecombinantglycoproteinsbymammaliancellcultures.Bothα(1-3)-linkedgalactoseandthelessabundantα(1-6)-linkedgalactosearecleavedbythisenzyme.Otherworkershaveusedtheenzymeforthemodificationofbloodgroupspecificity.Theenzymealsohasdiagnosticapplicationsforthedeterminationofraffinoseinfood.

SuggestionsforUse:
ProcedureForDe-galactosylation
1.Addupto100μgofglycoproteinor3nmolofoligosaccharidetotube.
2.Addde-ionizedwatertoatotalof14μl.
3.Add4μlof5xReactionBuffer.
4.Add2µlcontaining5mUα(1-3,4,6)-Galactosidase.
5.Incubateat37°Cfor18hours.

Note:Theenzymeisstable.At37°CinpH6.0buffer,fullactivityisrecoveredafter19hoursand70%after48hours.

Note:Theenzymehasaslowturnover.After18hoursat37°Cwith5U/mlenzyme,a40µMsolutionofasialo,biantennary,fucosylatedoligosaccharidewithouterarmα(1-3)-galactoseis67%de-α-galactosylated.

Shipswith:
WS00615xReactionBuffer[500mMsodiumcitrate/phosphate(pH6.0)]

ReactionBuffer:5Xconcentratedbufferwhichwhendilutedgives100mMsodiumcitrate-phosphatepH6.0.

Formulation:
Lyophilizedfrom100mMsodiumphosphatepH6.5,containing0.25mg/mlbovineserumalbumin.SeenoteaboveonBSAamount.

MW:~26kD

UnitDefinition:
Oneunitisdefinedastheamountofenzymerequiredtohydrolyze1μmoleofpNP-α-D-galactopyranosideperminuteatpH6.5and37oC.

Size:5Ulyophilized

ProductCode:GKX-5007