Purification and Characterization of Thermostable and Detergent-Stable Alpha-Amylase from Anoxybacillus sp. AH1
Year: 2015
Type: article
Abstract: A thermostable and detergent-stable α-amylase from a newly isolated Anoxybacillus sp.AH1 was purifi ed and characterized.Maximum enzyme production (1874.8U/mL) was obtained at 24 h of incubation.The amylase was purifi ed by using Sephadex G-75 gel fi ltration, aft er which an 18-fold increase in specifi c activity and a yield of 9 % were achieved.The molecular mass of the purifi ed enzyme was estimated at 85 kDa by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE).The optimum pH and temperature values of the enzyme were 7.0 and 60 °C, respectively.The enzyme was highly stable in the presence of 30 % glycerol, retaining 85 % of its original activity at 60 °C within 120 min.K m and v max values were 0.102 μmol and 0.929 μmol/min, respectively, using Lineweaver-Burk plot.The enzyme activity was increased by various detergents, but it was signifi cantly inhibited in the presence of urea.Mg 2+ and Ca 2+ also signifi cantly activated α-amylase, while Zn 2+ , Cu 2+ and metal ion chelators ethylenediaminetetraacetic acid (EDTA) and 1,10-phenanthroline (phen) greatly inhibited the enzyme activity.α-Amylase activity was enhanced by β-mercaptoethanol (β-ME) and dithiothreitol (DTT) to a great extent, but inhibited by p-chloromercuribenzoic acid (PCMB).Iodoacetamide (IAA) and N-ethylmaleimide (NEM) had a slight, whereas phenylmethylsulfonyl fl uoride (PMSF) had a strong inhibitory eff ect on the amylase activity.
Institution: Dicle University
Language: English
Subfield: Biotechnology
Domain: Life Sciences
Open Access status: diamond
Funder: Dicle Üniversitesi
OpenAlex ID: W4233152258