Exploration and functional expression of homologous lipases of Candida antarctica lipase B.

Candida (also known as Pseudozyma ) antarctica lipase B (CAL-B) has been intensely studied in academic and industrial fields. However, the research related to its homologous enzymes has been rarely reported. In the current investigation, protein sequenc e similarity search of CAL-B has been conducted and six homologous protein sequences were identified. After the syntheses of their codon-opt imized genes, the synthetic genes have been cloned into a periplasmic expression vector to express in Escherichia coli . Among six homologous sequences, four sequences were successfully expressed in E. coli . The hydrolytic activities of the expressed proteins towards 4-nitrophenyl acetate and 4-nitrophenyl butyrate were measured and compared with those of CAL-B to identify whether the expressed proteins wo rk as a hydrolase. It has been revealed that the expressed proteins can hydrolyze the substrates and the specific activities were deter mined as (1.3 – 30) × 10 -2 μ mol/min/mg, which are lower than those of CAL-B. Among these homologous enzymes, Pseudozyma hubeiensis SY62 exhibits the comparable enantioselectivity to that of CAL-B towards the hydrolysis of (±)-1-phenylethyl acetate.