食品科学2024，Vol.45Issue(18)：77-86,10.DOI:10.7506/spkx1002-6630-20240222-109
蛋白质分子表面氨基酸突变提高植酸酶YiAPPA的活性和热稳定性
Mutation of Amino Acid Residues on the Protein Surface Improves the Activity and Thermostability of Phytase YiAPPA
摘要
Abstract
In order to improve the activity and thermal stability of phytase and to enhance its application potential in the food processing industry,single-site mutants were constructed by homology modeling of phytase YiAPPA and directed mutagenesis of lysine(Lys)and glycine(Gly)residues located on the molecular surface.The mutant K216R with significantly improved activity and thermostability and the mutant K189R with improved thermostability were obtained.Subsequently,the combined mutant K189R/K216R was constructed and its enzyme activity and thermal stability were characterized.Compared with YiAPPA,the half-life of K189R/K216R at 80℃was increased from 14.81 to 23.35 min,the half-inactivation temperature(T5030)from 55.12 to 62.44℃,the melting temperature(Tm)from 48.36 to 53.18℃,and the specific enzymatic activity of K189R/K216R at 37℃and pH 4.5 from 3 959.98 to 4 469.13 U/mg.Molecular structure modeling analysis and molecular dynamics simulation revealed that the introduction of new hydrogen bonds into K189R/K216R could improve the stability of some structural units of the enzyme,thereby enhancing its thermostability.The improvement in its activity was mainly attributed to the increase in the volume of the catalytic pocket of K189R/K216R.In conclusion,the present study demonstrated that the strategy of amino acid residue mutation on the protein surface effectively improves the activity and thermostability of phytase YiAPPA.Our findings can serve as a reference for molecular modification of phytase and other enzymes.关键词
植酸酶/表面氨基酸/活性/热稳定性/分子动力学模拟Key words
phytase/amino acid residues located on the protein surface/activity/thermostability/molecular dynamics simulation分类
生物工程引用本文复制引用
曾静,郭建军,王通,袁林..蛋白质分子表面氨基酸突变提高植酸酶YiAPPA的活性和热稳定性[J].食品科学,2024,45(18):77-86,10.基金项目
国家自然科学基金地区科学基金项目(32160579) （32160579）
江西省主要学科学术和技术带头人培养计划项目(20212BCJ23033 （）
S2021GDQN2403) （）
江西省科学院基础研究项目(2022YJC2006) （2022YJC2006）
