Screening, Isolation, and Enzyme Kinetics of Bacterial Amylase collected from Rhizosphere soil

Abstract

Rhizosphere is a region where microbial communities are in complex association with the roots of plants where the activity of microbes and their enzymes are greatly influenced by root exudates. Amylase enzyme has great importance in biotechnology, with enormous utilization in food, fermentation, textile, and paper industries. They are produced intracellularly and extracellularly by different life forms including microorganisms. Microbial amylases are preferred over other sources because of their vast availability and it also meets the growing needs of industry. The present investigation deals with the isolation, screening, and enzyme kinetics of bacterial amylase from the rhizosphere soil samples collected from a fertile field. Soil samples were collected from the rhizosphere and amylase-producing bacteria screening was carried out by using a starch agar plate. Extracellular amylase was extracted from fermentation broth followed by quantification by starch-iodine assay. Bacterial amylase enzyme kinetics were determined by changing enzyme/substrate concentrations and incubation time. We successfully screened and isolated out starch hydrolyzing colonies from the rhizosphere soil samples. Studies on enzyme kinetics indicate that the activity of amylase increased initially as substrate and enzyme concentrations increased. If we kept enzyme concentration constant, to a certain point, there was no change in enzyme activity as the enzyme was saturated and no more enzyme was available to react with the excess substrate. Initially, enzyme activity increased as enzyme volume increased, but since substrate concentration was kept constant, higher volumes of the enzyme could not speed up the reaction. Further, under a prolonged incubation period, less amount of substrate was available at the end of a reaction. Therefore, it is concluded that the reaction velocity increases.