Cloning, Expressing And Application Of β-Dehalogenase

3-Hydroxypropionic acid is one of the top 12 platform chemicals in the word. We have screened out a Bacillus sp. CGMCC No.4196 strain from soli, which produced P-dehalogenase. This strain converted 3-chloroproionic into 3-hydroxypropionic acid. In this paper, we constructed a genomic library of Bacillus sp., and screened out theβ-dehalogenase gene. This 465bp gene was cloned into pET30a(+) plasmid. and transformed into Escherichia coli BL21(DE3)-CondonPlus. The recombinant enzyme was purified by HisTrapTMFF affinity chromatography. SDS-PAGE showed that the P-dehalogenase had a molecular weight of about 23.1 kDa. Further study on the enzymatic characteristics showed that with 3-chloroproionic as substrate, the optimum temperature and pH for the hydrolytic activity of rBhd were 30℃and 7.0, respectively. Km and Vmax were 3.26μ.mol/L and 17.86μmol/(mL·min). respectively.We have built up a P-dehalogenase conversion way for producing 3-hydroxypropionic acid from 3-chloroproionic. Preparing 3-hydroxypropionic with rBhd needed a moderate condition and had a high conversion ratio with no by-products. Using 10 mmol/L 3-chloroproionic as substrate, the conversion ratio was 93% in 32 h. The final concertration of 3-hydroxypropionic acid was about 9.4 mmol/L. This study provides groundwork for producing 3-hydroxypropionic acid by enzymes conversion methods.