化工进展2024，Vol.43Issue(3)：1466-1473,8.DOI:10.16085/j.issn.1000-6613.2023-0368
枯草芽孢杆菌表面展示氨肽酶的构建及协同水解大豆蛋白
Construction of aminopeptidase on the surface of Bacillus subtilis and its synergic hydrolysis of soybean protein
摘要
Abstract
Aminopeptidase is an important synergistic enzyme for protein deep hydrolysis,however the high cost and poor stability of free enzyme remain the primary limiting factors for their industrial applications.In this study,we constructed a new easily isolated and well tolerated whole-cell biocatalyst which displayed the aminopeptidase derived from Pseudomonas Aeruginosa GF31 on the surface of Bacillus subtilis WB800N spores by surface-display technology.And immunofluorescence analysis indicated that the aminopeptidase was correctly expressed on the surface of spores.The enzyme activity of surface-displayed aminopeptidase reached 75.61U/g at 60℃and pH=9.0.The aminopeptidase and alkaline protease were used in a synergistic hydrolysis of soybean protein,in which 5.0%of soybean protein was hydrolyzed for 8h at 60℃with an enzyme ratio of 2∶1 between the alkaline protease and the aminopeptidase,with an initial hydrolysis pH of 10.0 followed by 9.0.The maximum hydrolysis degree was 55.50%,which was 3.3-fold and 1.5-fold times higher than that achieved by surface-displayed aminopeptidase or alkaline protease alone,respectively.Simultaneously,the content of 16 kinds of free amino acids was significantly increased.Specifically,the content of hydrophobic amino acids including leucine,tyrosine,and phenylalanine increased by 19.21mg/L,8.59mg/L and 16.77mg/L,respectively.While the content of umami amino acid including glutamic acid and aspartic acid increased by 13.98mg/L and 4.11mg/L,respectively.These results suggested that aminopeptidase in deep hydrolysis could significantly reduce the bitterness of protein hydrolysate and enhance its umami favor,and had excellent potential for practical use.关键词
氨肽酶/表面展示/枯草芽孢杆菌/大豆蛋白/协同水解Key words
aminopeptidase/surface-display/Bacillus subtilis/soybean protein/synergistic hydrolysis分类
生物科学引用本文复制引用
袁晓黎,李青云,刘幽燕,李进,白雪,唐爱星..枯草芽孢杆菌表面展示氨肽酶的构建及协同水解大豆蛋白[J].化工进展,2024,43(3):1466-1473,8.基金项目
国家自然科学基金(21966007) （21966007）
广西自然科学基金(2018GXNSFAA281278). （2018GXNSFAA281278）
