Co2+-mediated time- and temperature-dependent activation of neutral alpha-D-mannosidase from monkey brain.
Neutral alpha-D-mannosidase from monkey brain was purified by Co2+-chelate affinity chromatography and immunoadsorbent affinity chromatography. The purified enzyme, with subunit Mr 45,000, was essentially homogeneous with only traces of two contaminant proteins as revealed by SDS/polyacrylamide-gel electrophoresis and AgNO3 staining. The purified enzyme, on preincubation with Co2+ at 37 degrees C or 60 degrees C followed by assay, showed a time-dependent enhancement in activity. The enhanced activity of the enzyme persisted even after removal of the Co2+. Bacitracin could partially prevent the activation. An aminopeptidase activity that was stimulated by Co2+ both at 37 degrees C and at 60 degrees C was present in the purified enzyme. After preincubation of the enzyme with Co2+ there was evidence for the release of amino acids, as revealed by t.l.c., but the Mr determined by SDS/polyacrylamide-gel electrophoresis was not appreciably altered. It is suggested that a Co2+-stimulated thermostable aminopeptidase, inseparable from the neutral mannosidase, may be involved in the stimulation of neutral mannosidase activity during its preincubation with Co2+.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001921:Brain / Q000201:enzymology*; D002846:Chromatography, Affinity; D002855:Chromatography, Thin Layer; D003035:Cobalt / Q000494:pharmacology*; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation / Q000187:drug effects; D005944:Glucosamine / Q000494:pharmacology; D000882:Haplorhini; D006863:Hydrogen-Ion Concentration; D007163:Immunosorbent Techniques; D008361:Mannosidases / Q000378:metabolism*; D013696:Temperature; D043323:alpha-Mannosidase
D000596:Amino Acids; D003035:Cobalt; D008361:Mannosidases; D043323:alpha-Mannosidase; D005944:Glucosamine