Abstract
Two novel genes (pwtsB and pwtsC) encoding lipases were isolated by screening the soil metagenomic library. Sequence analysis revealed that pwtsB encodes a protein of 301 amino acids with a predicted molecular weight of 33 kDa, and pwtsC encodes a protein of 323 amino acids with a predicted molecular weight of 35 kDa. Furthermore, both genes were cloned and expressed in Escherichia coli BL21 (DE3) using pET expression system. The expressed recombinant enzymes were purified by Ni-nitrilotriacetic acid affinity chromatography and characterized by spectrophotometric with different p-nitrophenyl esters. The results showed that PWTSB displayed a high degree of activity and stability at 20°C with an optimal pH of around 8.0, and PWTSC at 40°C with an optimal pH of around 7.0. P-nitrophenyl palmitate (p-NPP) was identified as the best substrate of PWTSB and PWTSC. The specific activities of PWTSB and PWTSC were 150 and 166 U/mg, respectively toward p-NPP at 30°C, about 20-fold higher than that toward p-nitrophenyl butyrate (C4) and caprylate (C8). In conclusion, our results suggest that PWTSB is a cold adapt lipase and PWTSC is a thermostable lipase to long-chain p-nitrophenyl esters.
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This work was supported by grant from Department of Health of Shandong Province (2007HZ036).
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Communicated by Erko Stackebrandt.
P. Wei and L. Bai contributed equally to this work.
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Wei, P., Bai, L., Song, W. et al. Characterization of two soil metagenome-derived lipases with high specificity for p-nitrophenyl palmitate. Arch Microbiol 191, 233–240 (2009). https://doi.org/10.1007/s00203-008-0448-5
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DOI: https://doi.org/10.1007/s00203-008-0448-5