A rational design for improving the pepsin resistance of cellulase E4 isolated from T.fusca based on the evaluation of the transition complex and molecular structure
When feeding enzymes are consumed by livestock and poultry,the stomach is the first organ to come in contact with those enzymes.As a result,the feeding enzymes will face two major hazards in the stomach: acidolysis and enzymatic hydrolysis.Therefore,it is very important to improve the tolerance of feed enzymes to the gastric juice environment.However,because the pepsin hydrolysis of amino acids is not specific,it is a very challenging problem to improve the resistance to pepsin through the mutation of amino acids in protein molecules.In this paper,we use protein docking technology(ZDOCK)to simulate the transition state of pepsin and E4 in the catalytic reaction process,decipher the key amino acid interactions on the surface of the transition state complex.Then,the key amino acids are mutated into non-preferred amino acids.At the same time,the molecular dynamics simulation was used to evaluate the stability of the mutant.Finally,we tested three optimal mutants(E4L691H,E4L691C,L800R and E4L691H,L800H)in a biological experiment,and the results showed that the half-life of the E4L691C,L800R protein was 60 times longer than that of the wild type,and the enzymatic properties were consistent with those of the wild type.This study provides a useful molecular improvement strategy for protein pepsin resistance.
Cellulase E4 Transition state Protease resistance Rational design Molecular simulation
Ying Wang Yanxia Zhou Gen Lu Daling Liu Chunfang Xie Dongsheng Yao
National Engineering Research Center of Genetic Medicine,Guangzhou City,Guangdong Province 510632 Ch Department of Bioengineering,Jinan University,Guangzhou City,Guangdong Province 510632 China;Institu
国内会议
第九届国际分子模拟与信息技术应用学术会议(ICMS&I2018)
太原
英文
356-373
2018-05-01（万方平台首次上网日期，不代表论文的发表时间）