Extracellular acid protease of Aspergillus oryzae grown on liquid media: multiple forms due to association with heterogeneous polysaccharides.
The acid protease (EC 2.4.23.6) that is produced extracellularly when Aspergillus oryzae is grown on liquid media has been isolated and characterized. The enzyme was purified by precipitation with tannic acid, chromatography on Duolite A-2, and gel filtration on Sephadex G-100. The last step yielded four active components, with varying molecular weights ranging from 42,000 to 60,000. Two of them, designated E1 and E1a, with molecular weights of 60,000 and 55,000, respectively, were heterogeneous on isoelectric focusing, both giving at least three enzyme species with different isoelectric points, whereas the other two, E1b and E2, with molecular weights of 49,000 and 42,000, respectively, were essentially homogeneous. These four enzymes activated bovine pancreatic trypsinogen and had the same pH optima in the acid pH range. They had essentially the same amino acid composition and immunologically cross-reacted with each other. These catalytic, chemical, and immunological properties are similar to those of acid protease A1 and A2 from A. oryzae grown on solid bran media. Unlike acid protease from solid bran culture, which contains both carbohydrate-containing and the carbohydrate-free species, all of the four enzymes, E1, E1a, E1b, and E2, contained carbohydrate, ranging from 18.9 to 43% and comprising three hexoses, glucose, galactose, and mannose. The carbohydrate portions were polysaccharide in nature and heterogeneous with respect to both molecular weight and sugar composition, and at least a part of the carbohydrate was present in the form of homopolysaccharides such as galactan and mannan. These findings indicate that polysaccharide chains with different molecular weights and with different chemical compositions are apparently responsible for the microheterogeneity of acid protease.
D000596:Amino Acids / Q000032:analysis; D000941:Antigens; D001230:Aspergillus / Q000201:enzymology*; D001236:Aspergillus oryzae / Q000201:enzymology*; D002241:Carbohydrates / Q000032:analysis; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D007527:Isoenzymes / Q000032:analysis* / Q000276:immunology / Q000378:metabolism; D008970:Molecular Weight; D010447:Peptide Hydrolases / Q000032:analysis* / Q000276:immunology / Q000378:metabolism; D011134:Polysaccharides / Q000032:analysis*
D000596:Amino Acids; D000941:Antigens; D002241:Carbohydrates; D007527:Isoenzymes; D011134:Polysaccharides; D010447:Peptide Hydrolases