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Detergent-resistant α-amylase derived from Anoxybacillus karvacharensis K1 and its production based on whey
Citation Link: https://doi.org/10.15480/882.13064
Publikationstyp
Journal Article
Date Issued
2024-12-01
Sprache
English
Author(s)
Ghevondyan, Diana
Soghomonyan, Tigran
Hovhannisyan, Pargev
TORE-DOI
Journal
Volume
14
Issue
1
Article Number
12682
Citation
Scientific Reports 14 (1): 12682 (2024)
Publisher DOI
Scopus ID
Publisher
Nature Publishing Group
In the field of biotechnology, the utilization of agro-industrial waste for generating high-value products, such as microbial biomass and enzymes, holds significant importance. This study aimed to produce recombinant α-amylase from Anoxybacillus karvacharensis strain K1, utilizing whey as an useful growth medium. The purified hexahistidine-tagged α-amylase exhibited remarkable homogeneity, boasting a specific activity of 1069.2 U mg−1. The enzyme displayed its peak activity at 55 °C and pH 6.5, retaining approximately 70% of its activity even after 3 h of incubation at 55 °C. Its molecular weight, as determined via SDS-PAGE, was approximately 69 kDa. The α-amylase demonstrated high activity against wheat starch (1648.8 ± 16.8 U mg−1) while exhibiting comparatively lower activity towards cyclodextrins and amylose (≤ 200.2 ± 16.2 U mg−1). It exhibited exceptional tolerance to salt, withstanding concentrations of up to 2.5 M. Interestingly, metal ions and detergents such as sodium dodecyl sulfate (SDS), Triton 100, Triton 40, and Tween 80, 5,5ʹ-dithio-bis-[2-nitrobenzoic acid (DNTB), β-mercaptoethanol (ME), and dithiothreitol (DTT) had no significant inhibitory effect on the enzyme’s activity, and the presence of CaCl2 (2 mM) even led to a slight activation of the recombinant enzyme (1.4 times). The Michaelis constant (Km) and maximum reaction rate (Vmax), were determined using soluble starch as a substrate, yielding values of 1.2 ± 0.19 mg mL−1 and 1580.3 ± 183.7 μmol mg−1 protein min−1, respectively. Notably, the most favorable conditions for biomass and recombinant α-amylase production were achieved through the treatment of acid whey with β-glucosidase for 24 h.
Subjects
Acid whey
Anoxybacillus karvacharensis
Biochemical characterization
Cloning
Expression
α-amylase
DDC Class
572: Biochemistry
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