Production and characterization of thermostable [alpha]-amylase from a newly isolated strain of Bacillus subtilis KIBGE-HAR

Investigation on the fermentation conditions for Alpha-amylase (1,4-[alpha]-D-glucan glucanohydrolase, E.C. 3.2.1.1) production was carried out with Bacillus subtilis KIBGE-HAR and a optimal synthetic medium for enzyme production was developed. Alpha-amylase production and cell population reached maximum after 24 hours of cultivation. The optimum temperature and pH for enzyme production were found to be 50[degrees]C and 7.0 respectively. Starch (15 g/l), which was used as a carbon source, supported the maximum production of enzyme. Peptone was used as a nitrogen source and the best concentration of peptone for [alpha]-amylase formation was found to be 5 g/l. High [alpha]-amylase titre was obtained in medium supplemented with 1.0 g/l yeast extract. Calcium chloride was added in the medium as a stabilizer and 0.2 mg/dl Ca[Cl.sub.2] was found to be the most favorable concentration for [??]-amylase production and stability. Among different reference media tested for [??]-amylase production, medium 3 gave the maximum yield of enzyme and our own optimized medium 4 was proved to be optimal for [??]-amylase production in contrast with the reference media. The optimum temperature and pH of the enzyme were found to be 60[degrees]C and 7.0 respectively. The most suitable buffer system for pH maintenance was proved to be Tris-HCl buffer (50 mM). Velocity of reaction reached to maximum in the presence of 2% substrate i.e. starch. Keywords: Fermentation | thermostable | medium | optimization | alpha amylase Gene Bank Accession: EU819144 (16S rRNA gene sequencing for Bacillus subtilis KIBGE-HAR)
Introduction The [alpha]-amylase (E.C. 3.2.1.1) randomly hydrolyzes alpha 1,4 glucosidic linkages in starch, glycogen and related polysaccharides yielding dextrins, oligosaccharides, maltose and D-glucose (Takeshita et al., 1975). Bacterial [alpha]-amylases are [...]