title: Purification and Characterization of Thermostable and Detergent-Stable α-Amylase fromAnoxybacillussp. AH1.
abstract: A thermostable and detergent-stable α-amylase from a newly isolatedAnoxybacillussp. AH1 was purified and characterized. Maximum enzyme production (1874.8 U/mL) was obtained at 24 h of incubation. The amylase was purified by using Sephadex G-75 gel filtration, after which an 18-fold increase in specific activity and a yield of 9% were achieved. The molecular mass of the purified 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.Kmandvmaxvalues 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 significantly inhibited in the presence of urea. Mg2+and Ca2+also significantly activated α-amylase, while Zn2+, Cu2+and metal ion chelators ethylenediaminetetraaceticacid(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 byp-chloromercuribenzoic acid (PCMB). Iodoacetamide (IAA) andN-ethylmaleimide (NEM) had a slight, whereas phenylmethylsulfonyl fluoride (PMSF) had a strong inhibitory effect on the amylase activity.
authors.affiliation: Molecular Biology and Genetic Department, Faculty of Science, Dicle University, TR-21280 Diyarbakır, Turkey.
authors.forename: Ömer
authors.initials: Ö
authors.lastname: Acer
authors.affiliation: Molecular Biology and Genetic Department, Faculty of Science, Dicle University, TR-21280 Diyarbakır, Turkey.
authors.forename: Fatma Matpan
authors.initials: FM
authors.lastname: Bekler
authors.affiliation: Molecular Biology and Genetic Department, Faculty of Science, Dicle University, TR-21280 Diyarbakır, Turkey.
authors.forename: Hemşe
authors.initials: H
authors.lastname: Pirinççioğlu
authors.affiliation: Division of Science Teaching, Ziya Gökalp Faculty of Education, Dicle University, TR-21280 Diyarbakır, Turkey.
authors.forename: Reyhan Gül
authors.initials: RG
authors.lastname: Güven
authors.affiliation: Molecular Biology and Genetic Department, Faculty of Science, Dicle University, TR-21280 Diyarbakır, Turkey.
authors.forename: Kemal
authors.initials: K
authors.lastname: Güven
keywords: Anoxybacillus sp. AH1; detergent-stable α-amylase; enzyme activity inhibition; enzyme purification