Purification and characterization of a Coffea canephora alpha-D-galactosidase isozyme.
Exoglycosidases modify carbohydrate epitopes on glycoproteins and glycolipids. The alpha-D-galactosidase from Coffea canephora is an important exoglycosidase which degrades the human blood group B epitope. Although multiple isozymes have been described, they have never been demonstrably purified and thoroughly characterized. We have developed a technique to purify an isozyme to homogeneity. The isolated enzyme has a molecular weight of 36.7 kDa by SDS PAGE and 34.0 kDa by gel filtration. The isozyme is highly selective for alpha-D-galactosides and inactive against other low molecular weight substrates. It hydrolyzes the the terminal alpha-D-galactosyl residue from the blood group B epitope. Protease activity is below detectable limits. The isozyme has a broad pH optima at 6.3, a pl of 7.03, is unaffected by ionic strength, and is stable at 4 degrees C.
Haibach
F
F
Department of Pathology, University of Missouri, Columbia.
Hata
J
J
Mitra
M
M
Dhar
M
M
Harmata
M
M
Sun
P
P
Smith
D
D
D000596:Amino Acids / Q000032:analysis; D002846:Chromatography, Affinity; D002850:Chromatography, Gel / Q000379:methods; D003069:Coffee / Q000201:enzymology*; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D007527:Isoenzymes / Q000302:isolation & purification* / Q000378:metabolism; D007700:Kinetics; D008970:Molecular Weight; D009994:Osmolar Concentration; D013379:Substrate Specificity; D000519:alpha-Galactosidase / Q000302:isolation & purification* / Q000378:metabolism
D000596:Amino Acids; D003069:Coffee; D007527:Isoenzymes; D000519:alpha-Galactosidase