Izvorni znanstveni članak
https://doi.org/10.17133/ftb.54.01.16.4122
Purification and Characterization of Thermostable and Detergent-Stable α-Amylase from Anoxybacillus sp. AH1
Ömer Acer
; Faculty of Science, Dicle University, Diyarbakır, Turkey
Fatma Matpan Bekler
; Faculty of Science, Dicle University, Diyarbakır, Turkey
Hemşe Pirinççioğlu
; Faculty of Science, Dicle University, Diyarbakır, Turkey
Reyhan Gül Güven
; Ziya Gökalp Faculty of Education, Dicle University, Diyarbakır, Turkey
Kemal Güven
; Faculty of Science, Dicle University, Diyarbakır, Turkey
Sažetak
A thermostable and detergent-stable α-amylase from a newly isolated Anoxybacillus sp. AH1 was purifi ed 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. Km and vmax values were 0.102 μmol and 0.929 μmol/min, espectively, 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 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 fluoride (PMSF) had a strong inhibitory effect on the amylase activity.
Ključne riječi
detergent-stable α-amylase; Anoxybacillus sp. AH1; enzyme activity inhibition; enzyme purification
Hrčak ID:
155098
URI
Datum izdavanja:
31.3.2016.
Posjeta: 1.417 *