Enhanced Production, Enzymatic Activity, and Thermostability of an α‑Amylase from Bacillus amyloliquefaciens in Lactococcus lactis

A novel α-amylase gene (BAA) from Bacillus amyloliquefaciens was cloned into Lactococcus lactis, designing two recombinant α-amylases to facilitate extracellular secretion. Following optimizing the expression conditions, the highest yield of BAA (88.12 mmol/L) was achieved upon 36 h induction and 5...

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Bibliographic Details
Published in:Journal of agricultural and food chemistry 2024-11, Vol.72 (44), p.24587-24598
Main Authors: Wei, Dehua, Ma, Tiange, Montalbán-López, Manuel, Li, Xingjiang, Wu, Xuefeng, Mu, Dongdong
Format: Article
Language:English
Subjects:
alpha-Amylases - chemistry
alpha-Amylases - genetics
alpha-Amylases - metabolism
Bacillus amyloliquefaciens - chemistry
Bacillus amyloliquefaciens - enzymology
Bacillus amyloliquefaciens - genetics
Bacillus amyloliquefaciens - metabolism
Bacterial Proteins - chemistry
Bacterial Proteins - genetics
Bacterial Proteins - metabolism
Biotechnology and Biological Transformations
Enzyme Stability
Hot Temperature
Hydrolysis
Lactococcus lactis - enzymology
Lactococcus lactis - genetics
Lactococcus lactis - metabolism
Starch - chemistry
Starch - metabolism
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Summary:A novel α-amylase gene (BAA) from Bacillus amyloliquefaciens was cloned into Lactococcus lactis, designing two recombinant α-amylases to facilitate extracellular secretion. Following optimizing the expression conditions, the highest yield of BAA (88.12 mmol/L) was achieved upon 36 h induction and 5 ng/mL nisin concentration. Determining the enzymatic properties of BAA revealed its poor stability and activity at high temperatures, hindering its widespread application. Therefore, we used computer-aided design to generate a mutant, S275L, which exhibited significantly improved activity and thermostability: an 18.7% increase in enzymatic activity (3767.38 U/mg), a 10 °C increase in optimal temperature, and a 49.2% improvement in stability at 60 °C. Molecular dynamics simulations and force analysis confirmed these enhancements. Finally, the mutant S275L’s potential was further analyzed for starch hydrolysis on poultry feed. Therefore, the mutant S275L holds promising as an enzyme agent for enhanced feed digestibility and quality.
ISSN:0021-8561
1520-5118
1520-5118
DOI:10.1021/acs.jafc.4c05070