Abstract
The product YkoN of the gene of unknown function, ykoN, of Bacillus subtilis Marburg has the pentapeptide lipase/esterase motif (Gly-X-Ser-X-Gly), and thus YkoN is expected to have a lipase or esterase activity. To characterize the expected enzyme activity the plasmid having a modified ykoN that include the sequence for His(x6) tag at its C-terminus of YkoN, which has 373 amino acid residues, was constructed. His-tagged YkoN protein of 39 kDa was induced in Escherichia coli BL21 (DE3) cells harboring chaperon plasmid pGro7 and purified to near homogeneity by using gel filtration and Ni-agarose. When p-nitrophenyl-esters of different fatty acid chain length were examined, the purified YkoN hydrolyzed the esters of fatty acid with short chain length (4-6 carbon atoms) preferentially. The esters of fatty acid with longer chain (C ≥ 10) were hydrolyzed inefficiently. The activity required no divalent cations and was not affected by addition of EDTA. The optimal pH for the activity was from pH 7.4 to pH 8.6. These results indicate that YkoN is a novel esterase which hydrolyzes the esters of fatty acid with short chain length.
