Improving the expression yield of Candida antarctica lipase B in Escherichia coli by mutagenesis.
Increasing the expression yield of active Candida antarctica lipase B (CAL-B) in Escherichia coli was achieved by using a codon-optimized synthetic gene and by mutagenesis to introduce hydrophilic residues on the surface of CAL-B. Five residues (four leucines and one isoleucine) on the surface of CAL-B were selected and changed with aspartate after codon optimization. While the codon-optimized synthetic gene of CAL-B did not increase the expression yield, the mutation increased the activity of the enzyme three-fold (3.3 mg/l of culture) compared to the wild type. The mutant enzyme had similar hydrolytic activity toward hydrolysis of p-nitrophenyl acetate or p-nitrophenyl butyrate and enantioselectivity toward hydrolysis of (R, S)-1-phenylethyl acetate compared to the wild-type enzyme.
Jung
Suhyun
S
Department of Chemistry, Center for NanoBio Applied Technology, College of Natural Sciences, Sungshin Women's University, Seoul 136-742, Korea.
Park
Seongsoon
S
D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D005656:Fungal Proteins; D057927:Hydrophobic and Hydrophilic Interactions; D008049:Lipase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D008958:Models, Molecular; D015394:Molecular Structure; D016297:Mutagenesis, Site-Directed / Q000379:methods*; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D009154:Mutation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D013379:Substrate Specificity
D005656:Fungal Proteins; D050505:Mutant Proteins; D008049:Lipase; C431349:lipase B, Candida antarctica