Journal
JOURNAL OF MOLECULAR MICROBIOLOGY AND BIOTECHNOLOGY
Volume 22, Issue 5, Pages 300-311Publisher
KARGER
DOI: 10.1159/000343819
Keywords
Functional screening; Gene cloning; Heterologous expression; Metagenomic library construction; Pichia pastoris
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Funding
- National Natural Science Fund [30800039]
- National Natural Science Foundation of China
- Eleven-Fifth Mega-Scientific Project on Infectious Diseases, China [2008ZX10003-005, 2008ZX10003-009]
- Institute of Pathogen Biology, Chinese Academy of Medical Sciences [2007IPB007]
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Lipases are important enzymes for various biotechnological applications. By using functional expression screening, lipZ03, a novel lipase gene, was isolated from a soil-derived metagenomic library. The gene was supposed to encode a protein of 617 amino acids with a C-terminal targeting signal region and four potential N-linked glycosylation sites. The protein sequence shared a conserved GXSXG motif (X represents any amino acid residue) with other microbial lipases. Gene lipZ03 was expressed in Pichia pastoris and the molecular weight was estimated to be approximately 65 kDa by electrophoresis. The optimum reaction temperature and pH value for LipZ03 was 50 degrees C and 9.0, respectively. The enzyme was highly stable in the temperature range of 40-60 degrees C and under alkaline conditions (pH 8-10). Lipolytic activity was significantly enhanced by Ca2+ and Mg2+ ions, but dramatically inhibited by Cu2+, Ni2+ and Hg2+ ions and EDTA. The purified enzyme preferentially hydrolyzed relatively long-chain triacylglycerols and was a true lipase rather than an esterase. Using a multi-stepwise methanol supply, the purified LipZ03 achieved a conversion yield of biodiesel production up to 74% after 36 h. Some interesting characteristics described here showed that the recombinant lipase may have potential to be a useful enzyme in industrial applications. Copyright (c) 2012 S. Karger AG, Basel
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