Purification and characterization of moderately thermostable raw-starch digesting α-amylase from endophytic Streptomyces mobaraensis DB13 associated with Costus speciosus

Endophytic actinobacteria are known to produce various enzymes with potential industrial applications. Alpha -amylase is an important class of industrial enzyme with a multi -dimensional utility. The present experiment was designed to characterize a moderately thermostable alpha -amylase producing endophytic Streptomyces mobaraensis DB13 isolated from Costus speciosus (J. Koenig) Sm. The enzyme was purified using 60% ammonium sulphate precipitation, dialysis, and Sephadex G-100 column chromatography. Based on 12% SDS-PAGE, the molecular weight of the purified alpha -amylase was estimated to be 55 kDa. The maximum alpha -amylase activity was achieved at pH 7.0, 50 degrees C and it retained 80% of its activity at both pH 7.0 and 8.0 after incubation for 2 h. The alpha -amylase activity is strongly enhanced by Ca 2+ , Mg 2+ , and inhibited by Ba 2+ . The activity remains stable in the presence of Tween-80, SDS, PMSF, and Triton X-100; however, beta -mercaptoethanol, EDTA, and H 2 O 2 reduced the activity. The kinetic parameters K m and V max values for this alpha -amylase were calculated as 2.53 mM and 29.42 U/mL respectively. The alpha -amylase had the ability to digest various raw starches at a concentration of 10 mg/mL at pH 7.0, 50 degrees C, where maize and rice are the preferred substrates. The digestion starts after 4 h of incubation, which reaches maximum after 48 h of incubation. These results suggest that S. mobaraensis DB13 is a potential source of moderately thermostable alpha -amylase enzyme, that effciently hydrolyzes raw starch. It suggesting that this alpha -amylase is a promising candidate to be use for industrial purposes.