title: Cloning and expression of a gene with phospholipase B activity from Pseudomonas fluorescens in Escherichia coli.
abstract: A gene from Pseudomonasfluorescens BIT-18 encoding a protein with phospholipase B activity (Pf-PLB) was cloned in E. coli BL21 (DE3). The open reading frame consists of 1272 bp and potentially encodes a protein of 423 amino acid residues with a calculated molecular mass of 45.8 kDa. The nucleotide sequence of Pf-PLB is 45%, 42%, 41%, 40%, 33%, and 31% identical to that of Bifidobacterium animals, Mycobacterium parascrofulaceum, Acidobacterium capsulatum, Lactobacillus johnsonii, Moraxella bovis, and Moraxella catarrhalis, respectively. The His-tagged protein was purified by affinity chromatography and the eluted protein hydrolyzed both the 1- and 2-ester bond of phosphatidylcholine. The recombinant Pf-PLB had optimal activity at pH 6.0 and 30 °C, and it showed 20.1% higher efficiency in the conversion rate of the phosphorus content than the wild-type.
authors.affiliation: School of Life Science, Beijing Institute of Technology, 100081 Beijing, PR China.
authors.forename: Fangyan
authors.initials: F
authors.lastname: Jiang
authors.forename: Shen
authors.initials: S
authors.lastname: Huang
authors.forename: Kaleem
authors.initials: K
authors.lastname: Imadad
authors.forename: Chun
authors.initials: C
authors.lastname: Li
mesh_terms: D003001:Cloning, Molecular / Q000379:methods; D004789:Enzyme Activation; D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology* / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology; D008245:Lysophospholipase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011551:Pseudomonas fluorescens / Q000201:enzymology* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D011994:Recombinant Proteins; D008245:Lysophospholipase