Preliminary characterization of a new exo-beta-(1,4)-galactanase with transferase activity.
As a prerequisite to the study of the fine chemical structure of the branched region of pectin, an exo-beta-(1,4)-galactanase was purified from a commercial preparation (Pectinex AR). Purification was carried out by precipitation with 70% saturated ammonium sulfate, preparative electrofocusing, anion-exchange chromatography and affinity chromatography on cross-linked alginate. Exogalactanase specific activity was 992 nkat mg-1 and the enzyme was devoid of beta-(1,3)- or beta-(1,6)-galactanase, arabinanase, beta-D-galactosidase and alpha-L-arabinofuranosidade activities. Residual exopolygalacturonase activity represented 2.9% of the galactanase activity. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and isoelectric focusing showed two close bands with molecular weights of 120,000 and 90,000 and pHi of 3.8 and 4.1, respectively. The enzyme acted in an exo manner and its activity was optimum at pH 3.5 and 60 degrees C. When incubated with galacto-oligosaccharides, new oligosaccharides with a higher degree of polymerization appeared, indicating the ability of the enzyme to transfer galactose residues.
Bonnin
E
E
Laboratoire de Biochimie et Technologie des Glucides, Institut National de la Recherche Agronomique, Nantes, France.
Lahaye
M
M
Vigouroux
J
J
Thibault
J F
JF
D001234:Aspergillus niger / Q000201:enzymology*; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D005656:Fungal Proteins / Q000302:isolation & purification* / Q000378:metabolism; D005685:Galactans / Q000378:metabolism; D005700:Galactosyltransferases / Q000302:isolation & purification* / Q000378:metabolism; D006026:Glycoside Hydrolases; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing; D008970:Molecular Weight; D009844:Oligosaccharides / Q000378:metabolism; D010368:Pectins / Q000378:metabolism; D013379:Substrate Specificity; D013696:Temperature; D001616:beta-Galactosidase / Q000302:isolation & purification* / Q000378:metabolism
D005656:Fungal Proteins; D005685:Galactans; D009844:Oligosaccharides; D010368:Pectins; D005700:Galactosyltransferases; D006026:Glycoside Hydrolases; C066350:exo-1,4-beta-galactanase; D001616:beta-Galactosidase