乳业科学与技术2025，Vol.48Issue(3)：11-17,7.DOI:10.7506/rykxyjs1671-5187-20241210-105
N81位点突变调控Ligilactobacillus cholophilus BD7642胆盐水解酶g1294特异性
Mutations at the N81 Site Modulate the Specificity of Bile Salt Hydrolase g1294 from Ligilactobacillus cholophilus BD7642
摘要
Abstract
This study aimed to analyze the effect of active site mutations on the specificity of bile salt hydrolase(BSH)g1294 from Ligilactobacillus cholophilus BD7642.Glycine scanning technology was used to investigate systematic mutations of BSH g1294's active center,and the activity of mutants was assessed by the spot-plate method.High performance liquid chromatography was utilized to analyze the specificity of the mutants toward six bile salts.Additionally,saturation mutagenesis was conducted at the N81 site,followed by molecular docking simulations to analyze the differences between the mutants and the wild type.Results revealed that mutations at some sites led to the inactivation of the BSH activity.However,the N81G and N174G mutants exhibited enhanced substrate specificity,the former being superior to the latter.Among the mutants at the N81 site,significant variations in the enzymatic activity and hydrolytic capability were observed.Molecular docking simulations indicated that the N81G mutant acquired hydrolytic capability by reducing the steric hindrance between the catalytic site Cys2 and glycocholic acid.In conclusion,this mutant showed improved specificity than did the wild type.The molecular recognition of bile acids by BSH may not be simply based on amino acid recognition,which could provide new insights for rational design for BSH engineering.关键词
Ligilactobacillus cholophilus/胆盐水解酶/特异性/异源表达/饱和突变Key words
Ligilactobacillus cholophilus/bile salt hydrolase/specificity/heterologous expression/saturation mutagenesis分类
生物科学引用本文复制引用
任全路..N81位点突变调控Ligilactobacillus cholophilus BD7642胆盐水解酶g1294特异性[J].乳业科学与技术,2025,48(3):11-17,7.基金项目
"十四五"国家重点研发计划重点专项(2022YFD2100704) （2022YFD2100704）
上海市国资委企业创新发展和能级提升项目(2022013) （2022013）
