Biochemical characterization of a family IV esterase with R-form enantioselectivity from a compost metagenomic library

A novel family IV esterase (hormone-sensitive lipase, HSL) gene, est15L, was isolated from a compost metagenomic library. Encoded Est15L comprised 328 amino acids with a molecular weight of 34,770 kDa and was an intracellular esterase without a signal peptide. The multiple sequence alignment (MSA) of Est15L with other family IV esterases showed conserved regions such as HGG, DYR, GXSXG, DPL, and GXIH. Native Est15L was a dimeric form from the results of size exclusion chromatography. It was optimally active at 50 ℃ and pH 9.0, indicating alkaline esterase. However, it showed a low thermostability with half-lives of 30.3 at 30 ℃ and 2.7 min at 40 ℃. It preferred p-nitrophenyl butyrate (C4) with Km and Vmax values of 0.28 mM and 270.8 U/mg, respectively. Est15L was inhibited by organic solvents such as 30% methanol, isopropanol, and acetonitrile with residual activities of 12.5, 0.9, and 0.3%, respectively. It was also inhibited by 1% SDS and 1% PMSF; however, Est15L maintained its activity at 1% Triton X-100 and EDTA. Est15L was inhibited by Cu2+, Zn2+, Mn2+, Co2+, Fe2+, and Na+. In addition, Est15L hydrolyzed glyceryl tributyrate with a residual substrate amount of 43.7% at 60 min but could not hydrolyze the oils (fish and olive) and glyceryl trioleate. Interestingly, Est15L showed significant enantioselectivity toward the R-form with a residual substrate amount of 44.6%, lower than that of the S-form (83.5%). Considering its properties, Est15L can be a potential candidate for chemical reactions, such as the synthesis of pharmaceutical compounds.