title: Cloning, expression, and characterization of a lipolytic enzyme gene (lip8) from Pseudomonas aeruginosa LST-03.
abstract: A lipolytic enzyme gene (lip8) was cloned from organic solvent-tolerant Pseudomonas aeruginosa LST-03 and sequenced. In the sequenced nucleotides, an open reading frame consisting of 1,173 nucleotides and encoding 391 amino acids was found. Lip8 is considered to belong to the family VIII of lipolytic enzymes whose serine in the consensus sequence of -Ser-Xaa-Xaa-Lys- acts as catalytic nucleophile. The gene was expressed in Escherichia coli and purified by a combination of ammonium sulfate fractionation and hydrophobic interaction and ion-exchange chromatographies to homogeneity on SDS-PAGE analysis. The optimum temperature and heat stability of Lip8 were not as high as those of Lip3 and LST-03 lipase, two other lipolytic enzymes from the same strain. Addition of glycerol to a solution containing Lip8 stabilized this enzyme. By measuring the activities against various triacylglycerols and fatty acid methyl esters having carbon chains of different lengths, Lip8 was categorized as an esterase which has higher activities against fatty acid methyl esters with short-chain fatty acids.
authors.affiliation: Department of Chemical Engineering, Osaka Prefecture University, Osaka, Japan. ogino@chemeng.osakafu-u.ac.jp
authors.forename: Hiroyasu
authors.initials: H
authors.lastname: Ogino
authors.forename: Takashi
authors.initials: T
authors.lastname: Mimitsuka
authors.forename: Toshiaki
authors.initials: T
authors.lastname: Muto
authors.forename: Miharu
authors.initials: M
authors.lastname: Matsumura
authors.forename: Masahiro
authors.initials: M
authors.lastname: Yasuda
authors.forename: Kosaku
authors.initials: K
authors.lastname: Ishimi
authors.forename: Haruo
authors.initials: H
authors.lastname: Ishikawa
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D020134:Catalytic Domain / Q000235:genetics; D005591:Chemical Fractionation; D002853:Chromatography, Liquid; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000737:chemistry / Q000302:isolation & purification; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005227:Fatty Acids / Q000378:metabolism; D005232:Fatty Acids, Volatile / Q000378:metabolism; D005798:Genes, Bacterial; D005990:Glycerol / Q000494:pharmacology; D008049:Lipase / Q000235:genetics* / Q000378:metabolism*; D008969:Molecular Sequence Data; D011550:Pseudomonas aeruginosa / Q000201:enzymology*; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D013379:Substrate Specificity; D013696:Temperature; D014280:Triglycerides / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D004269:DNA, Bacterial; D005227:Fatty Acids; D005232:Fatty Acids, Volatile; D011994:Recombinant Proteins; D014280:Triglycerides; D008049:Lipase; D005990:Glycerol