Extracellular Maltase of Bacillus brevis

Abstract

Bacillus brevis NRRL B-4389 produced extracellular maltase (α-glucosidase; EC 3.2.1.20) only in the presence of short α-1,4-glucosidic polymers, such as maltose and maltotriose. An optimum medium was developed; it contained 2.5% maltose, 0.5% nonfat dry milk, 0.4% yeast extract, and 0.01% CaCl 2 . The enzyme was produced extracellularly during the logarithmic phase of growth; no cell-bound activity was detected at any time. Partial purification of the maltase was accomplished by using diethylaminoethyl cellulose batch adsorption, ammonium sulfate precipitation, and Sephadex G-200 gel filtration. Maltase, isomaltase (oligo-1,6-glucosidase), and glucosyltransferase activities were purified 20.0-, 19.1-, and 11.5-fold, respectively. Some properties of the partially purified maltase were determined: optimum pH, 6.5; optimum temperature, 48 to 50°C; pH stability range, 5.0 to 7.0; temperature stability range, 0 to 50°C; isoelectric point, pH 5.2; and molecular weight, 52,000. The relative rates of hydrolysis of maltose (G 2 ), maltotriose (G 3 ), G 4 , methyl-α- d -maltoside, G 40 , dextrin, and isomaltose were 100, 22, 12, 10, 10, 8, and 5%, respectively; the K m on maltose was 5.8 mM; d -glucose, p -nitrophenyl-α- d -glucoside, and tris (hydroxymethyl) aminomethane were competitive inhibitors; transglucosylase activity of the enzyme on maltose resulted in the synthesis of isomaltose, isomaltotroise, and larger oligosaccharides.