Expression and identification of truncation Lactobacillus casei invertase based on rational design
In this study,a invertase-producing strain was screened and identified from soil samples of Qiandao Lake.The Lactobacillus casei invertase gene(LevH1)was amplified from L.casei complementary deoxyribonucleic acid(cDNA),and the recombinant L.casei invertase LEVH1-LC was ob-tained by constructing Escherichia coli.On the basis of rational design of enzyme,four invertase truncates LevH 1-LC-N,LevH 1-LC-C,LevH 1-LC-NC and LevH1-LC-M were obtained by gene truncation technique,and their enzymatic properties were studied.The results showed that an excellent in-vertase-producing strain LC23 was obtained,which was identified as L.casei with enzyme activity of 0.06 U/ml.The LevHl-LC enzyme activity of recombinant L.casei invertase was 8.37 U/ml,which was 138 times higher than that of natural invertase.Compared with recombinant L.casei inver-tase LevH1-LC,the enzyme activities of truncates LevH1-LC-N and LevH1-LC-NC were increased 4.1 times and 6.6 times,respectively,and the en-zyme activity was increased 568.5 times and 923.7 times compared with natural invertase.The results of enzyme kinetic analysis showed that the catalytic efficiency of truncate LevH 1-LC-NC was the highest,which was 29.7 times that of recombinant L.casei invertase LevH1-LC.The optimum reaction temperature and pH were 50 ℃ and 6.0,respectively,indicating good thermal stability and pH stability.Therefore,the truncate LevH1-LC-NC was a superior invertase mutant.
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