Analysis of the gene that regulate production of Vibrio cholerae El Tor cytolysin

从霍乱弧菌O1的Tn5插入突变体库中，我们发现了一个突变体NF404，该突变株即使在溶血素和霍乱毒素结构基因的外面也不能产生溶血素和霍乱毒素(CT)。对野生型霍乱弧菌的同源区进行克隆和测序，结果表明，该突变株编码39.5 kDa的蛋白，被命名为lypA。LypA基因推导的氨基酸序列与大肠杆菌溶血磷脂酶L2的同源性为37.6%。在NF404中引入LypA后，霍乱毒素和溶血素的产生丧失，溶血磷脂酶活性和CT产生恢复，但不能恢复溶血活性。单交叉插入灭活野生型菌株的LypA可降低溶血磷脂酶L2活性和霍乱毒素的产量，但不影响溶血活性。这些缺陷菌株降低了肠道毒性。这些结果表明，lypA可能是霍乱弧菌表达CT所必需的，并可能在霍乱弧菌的致病中起作用。

From the Tn5-inserted mutant library of Vibrio cholerae O1, we found a mutant, NF404, which lost the production of both hemolysin and cholera toxin (CT) even though the Tn5-insertion site was out side from the structural genes for hemolysin and CT.Cloning and sequencing analysis of the homologous region from the wild-type strain, revealed that the sequence spanning the coding region of an ORF1 nominated as lypA,encoding a 39.5 kDa protein. Deduced amino acid sequence of the lypA gene had 37.6% identity to the lysophospholipase L2 of Escherichia coli. Introduction of lypA into NF404 has lost cholera toxin and hemolysin productions, restored the lysophospholipase activity and CT production but not the hemolytic activity. Inactivation of lypA of the wild-type strains by single crossover insertion decreased the lysophospholipase L2 activity and cholera toxin production but not the hemolytic activity. These defective strains decreased enterotoxicity. These results suggest that lypA is possibly required for the expression of CT and may play a role in pathogenicity of Vibrio cholerae.