Cloning and expression of a gene with phospholipase B activity from Pseudomonas fluorescens in Escherichia coli
Cloning and expression of a gene with phospholipase B activity from Pseudomonas fluorescens in Escherichia coli
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DOI：
10.1016/j.biortech.2011.09.112
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发表时间：
2012-01-01
影响因子：
11.4
通讯作者：
Li, Chun
中科院分区：
文献类型：
--
作者：
Jiang, Fangyan;Huang, Shen;Li, Chun
从荧光假单胞菌（Pseudomonasfluorescens）BIT-18中克隆了一个编码具有磷脂酶B活性蛋白（Pf-PL B）的基因。coli BL21（DE3）。该基因的开放阅读框长1272 bp，编码423个氨基酸，分子量为45.8 kDa。Pf-PLB的核苷酸序列与动物双歧杆菌（Bifidobacterium animals）、副棒状分支杆菌（Mycobacterium parascrofulaceum）、荚膜酸杆菌（Acidobacterium capsulatum）、约氏乳杆菌（Lactobacillus johnsonii）、牛莫拉氏菌（Moraxella bovis）和卡他莫拉氏菌（Moraxella catarrhalis）的核苷酸序列的同源性分别为45%、42%、41%、40%、33%和31%。His标记的蛋白质通过亲和层析纯化，洗脱的蛋白质水解磷脂酰胆碱的1-和2-酯键。重组Pf-PLB在pH 6.0和30 ℃下具有最佳活性，磷含量转化率比野生型高20.1%。(C)2011爱思唯尔有限公司版权所有。
A gene from Pseudomonas fluorescens 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 degrees C, and it showed 20.1% higher efficiency in the conversion rate of the phosphorus content than the wild-type. (C) 2011 Elsevier Ltd. All rights reserved.