title: Isolation of a novel lipase gene from Serratia liquefaciens S33 DB-1, functional expression in Pichia pastoris and its properties.
abstract: A new lipase gene designated as SlLipA was isolated from Serratia liquefaciens S33 DB-1 by the genomic-walking method. The cloned gene contained an open reading frame (ORF) of 1,845 bp encoding 615 amino acids with a conserved GXSXG motif. Genome sequence analysis showed that an aldo/keto reductase gene closed to the SlLipA gene. The lipase gene was cloned into the expression vector pPICZalphaA and successfully integrated into the heterologous host, methylotrophic yeast Pichia pastoris GS115. Five transformants could be expressed as secreted recombinant proteins with the high activity on Triglyceride-Agarose plate and as candidates to produce the recombinant enzyme. A C-terminal His tag was used for its purification. The lipase activity of different transformants against substrate para-nitrophenyl laurate (p-NPL) varied from 14 to 16 U ml(-1). For the substrates para-nitrophenyl caprate (p-NPC), p-NPL, para-nitrophenyl myristate (p-NPM), para-nitrophenyl palmitate (p-NPP), and para-nitrophenyl stearate (p-NPS), the specific activity was shown to be preferred to long acyl chain length of p-NPS.
authors.affiliation: Plant Biotechnology Research Center, School of Agriculture and Biology, Shanghai Jiao Tong University, Shanghai, 200030, PR China.
authors.forename: Hongyan
authors.initials: H
authors.lastname: Yao
authors.forename: Shunwu
authors.initials: S
authors.lastname: Yu
authors.forename: Lida
authors.initials: L
authors.lastname: Zhang
authors.forename: Kaijing
authors.initials: K
authors.lastname: Zuo
authors.forename: Hua
authors.initials: H
authors.lastname: Ling
authors.forename: Fei
authors.initials: F
authors.lastname: Zhang
authors.forename: Kexuan
authors.initials: K
authors.lastname: Tang
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D017124:Conserved Sequence; D004591:Electrophoresis, Polyacrylamide Gel; D015971:Gene Expression Regulation, Enzymologic; D008049:Lipase / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D008969:Molecular Sequence Data; D010843:Pichia / Q000201:enzymology* / Q000235:genetics; D016415:Sequence Alignment; D044090:Serratia liquefaciens / Q000201:enzymology* / Q000235:genetics; D013379:Substrate Specificity; D019076:Transgenes / Q000235:genetics
chemical_list: D008049:Lipase