By the error-prone PCR and high throughput screening based on the color reaction of chloride ion concentration, a mutant library of R-2-CPA dehalogenase was created and screened. The specific activity of the mutants, called the DehDIV-G2 and DehDIV-E7, increased by 24.8% and 39.6%, respectively. Molecular docking of the enzymes with substrates was carried out with SYBYL. Docking scoring displayed that DehDIV-G2 decreased the activation energy of 0.961 4 kJ/mol,and DehDIV-E7 decreased the activation energy of 2. 549 8 kJ/mol. The mutant enzymes decreased the activation energy, and increased the affinity with R-2-CPA,thus the enzyme specific activities increased.%通过易错PCR手段将R-2-氯丙酸脱卤酶定向进化,并使用基于Cl-浓度显色反应的高通量筛选得到有效突变子库,发现突变子DehDIV-G2和DehDIV-E7的酶比活力分别提高25.2％和38.7％.通过SYBYL对酶与底物进行分子对接显示,DehDIV G2的活化能下降0.961 4 kJ/mol,DehDIV E7的活化能下降2.549 8 kJ/mol.由于酶和底物R-2-氯丙酸的活化能下降,亲和能力提高,从而提高酶的比活力.
展开▼
