Molecular and immunological characterization of OprL, the 18 kDa outer-membrane peptidoglycan-associated lipoprotein (PAL) of Pseudomonas aeruginosa
Molecular and immunological characterization of OprL, the 18 kDa outer-membrane peptidoglycan-associated lipoprotein (PAL) of Pseudomonas aeruginosa
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DOI：
10.1099/00221287-143-5-1709
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发表时间：
1997-05-01
期刊：
影响因子：
2.8
通讯作者：
Cornelis, P
中科院分区：
文献类型：
--
作者：
Lim, A;DeVos, D;Cornelis, P
对铜绿假单胞菌cosmid文库进行免疫学筛选，鉴定出产生18kda外膜蛋白的克隆。该蛋白与铜绿假单胞菌（P. aeruginosa）外膜蛋白的多克隆抗血清和OprL（肽聚糖相关外膜脂蛋白（PAL））的单克隆抗体（MA1-6）在Western blots中反应。对表达oprL的pOML7亚克隆进行测序，发现一个504 bp的ORF编码一个具有典型脂蛋白信号识别序列的多肽。在oprL上游发现另一个ORF，与大肠杆菌和流感嗜血杆菌的TolB蛋白同源。在oprL的下游，发现了另一个长321 bp的ORF (orf2)，编码一个带有信号肽的蛋白，与已知生物学功能的蛋白无同源性。在orf2的终止密码子之后，检测到一个rho无关的终止序列，该序列是P. aeruginosa PAO1插入元件1S222的一部分。OprL与革兰氏阴性菌的所有已知PALs具有同源性，特别是在c端部分。mAb MA1-6在免疫荧光中与铜绿假单胞菌细胞和表达oprL的大肠杆菌细胞反应，后者具有异常的细长形态，表明该蛋白的产生扰乱了分裂过程。
Immunological screening of a Pseudomonas aeruginosa cosmid library led to the identification of clones producing an 18 kDa outer-membrane protein. This protein reacted in Western blots with a polyclonal antiserum against outer-membrane proteins of P. aeruginosa and with a monoclonal antibody (MA1-6) specific for OprL, the peptidoglycan-associated outer-membrane lipoprotein (PAL). Sequencing of pOML7, a subclone expressing oprL, revealed an ORF of 504 bp encoding a polypeptide with a typical lipoprotein signal recognition sequence. Another ORF was found upstream of oprL, with homology to the TolB protein of Escherichia coli and Haemophilus influenzae. Downstream of oprL, a second ORF, of 321 bp, was found (orf2), encoding a protein with a signal peptide and with no homology with proteins of known biological function. After the stop codon of orf2, a rho-independent terminator sequence was detected which is part of the P. aeruginosa PAO1 insertion element 1S222. OprL showed homologies with all known PALs from Gram-negative bacteria, especially in the C-terminal part. mAb MA1-6 reacted with P. aeruginosa cells in immunofluorescence, and with E. coli cells expressing oprL, which had an abnormal, elongated morphology, an indication that production of the protein perturbed the division process.