title: Purification and some properties of three forms of glucoamylase from a Rhizopus species.
abstract: 1. Three forms of glucoamylase [EC 3.2.1.3] were simultaneously purified from a Rhizopus species by (NH4)2SO4 fractionation and successive chromatographies on Sephadex G-75, DEAE-Sephadex, and CM-Sephadex, and were finally separated from each other by means of recycling chromatography on Bio-Gel P-150. The purification achieved was 3--4 fold from crude extract with respect to each glucoamylase; the yields of the three glucoamylases, designated as Gluc1, Gluc2, and Gluc3 in order of content, were 39, 7, and 0.4%, respectively. All the purified enzymes were homogeneous in polyacrylamide gel electrophoresis, isoelectric focusing, and ultracentrifugation. 2. The three glucoamylases were glycoproteins differing in both amino acid composition and carbohydrate content, but showed a common antigenicity in immunodiffusion. The molecular weights of Gluc1, Gluc2, and Gluc3 were estimated to be 74,000, 58,600, and 61,400, respectively, by sedimentation equilibrium and these values were verified by SDS-polyacrylamide gel electrophoresis. The specific activities of the three enzymes toward starch were in the opposite order to their molecular weights. 3. The three glucoamylases had the same broad pH optima in the range pH 4.5--5.0 and shared a common susceptibility to inactivation by heat, extreme pH, and such divalent cations as Hg2+, Pb2+, and Mn2+, indicating close similarity in enzymatic properties.
mesh_terms: D000596:Amino Acids / Q000032:analysis; D002241:Carbohydrates / Q000032:analysis; D002413:Cations, Divalent / Q000494:pharmacology; D005087:Glucan 1,4-alpha-Glucosidase / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D005959:Glucosidases / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008970:Molecular Weight; D012233:Rhizopus / Q000201:enzymology*
chemical_list: D000596:Amino Acids; D002241:Carbohydrates; D002413:Cations, Divalent; D005959:Glucosidases; D005087:Glucan 1,4-alpha-Glucosidase