Endochitinase is transported to the extracellular milieu by the eps-encoded general secretory pathway of Vibrio cholerae
Endochitinase is transported to the extracellular milieu by the eps-encoded general secretory pathway of Vibrio cholerae
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DOI：
10.1128/jb.180.21.5591-5600.1998
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发表时间：
1998-11-01
影响因子：
3.2
通讯作者：
Folster, JP
中科院分区：
文献类型：
--
作者：
Connell, TD;Metzger, DJ;Folster, JP
霍乱弧菌的chiA基因编码降解几丁质的多肽，几丁质是在真菌的细胞壁和昆虫和甲壳类动物的外皮中发现的N-乙酰基-葡糖胺（GlcNAc）的均聚物。chiA具有对应于846个氨基酸的多肽的编码能力，其预测分子量为88.7kDa。在chiA开放阅读框的上游发现了具有启动子活性的52-bp区域。该基因的染色体拷贝的插入失活证实了霍乱弧菌几丁质酶活性的表达需要chiA。荧光类似物用于证明ChiA的酶活性对位于几丁质中GlcNAc单体之间的β，1 - 4糖苷键是特异性的。通过用MaIE-ChiA杂合蛋白免疫兔获得针对ChiA的抗体。与ChiA具有抗原相似性的多肽由霍乱弧菌的经典和E1 Tor生物型以及密切相关的细菌嗜水气单胞菌表达。使用野生型菌株569B和分泌突变体M14的免疫印迹实验证实了ChiA是由eps系统分泌的胞外蛋白。eps系统还负责分泌霍乱毒素，这是一种与ChiA没有氨基酸同源性的寡聚蛋白。这些结果表明，ChiA和霍乱毒素具有功能相似的胞外转运信号，这些信号对于eps依赖的分泌是必需的。
The chiA gene of Vibrio cholerae encodes a polypeptide which degrades chitin, a homopolymer of N-acetyl-glucosamine (GlcNAc) found in cell walls of fungi and in the integuments of insects and crustaceans. chiA has a coding capacity corresponding to a polypeptide of 846 amino acids having a predicted molecular mass of 88.7 kDa, A 52-bp region with promoter activity was found immediately upstream of the chiA open reading frame. Insertional inactivation of the chromosomal copy of the gene confirmed that expression of chitinase activity by V. cholerae required chiA. Fluorescent analogues were used to demonstrate that the enzymatic activity of ChiA was specific for beta,1-4 glycosidic bonds located between GlcNAc monomers in chitin, Antibodies against ChiA were obtained by immunization of a rabbit with a MaIE-ChiA hybrid protein. Polypeptides with antigenic similarity to ChiA were expressed by classical and E1 Tor biotypes of V. cholerae and by the closely related bacterium Aeromonas hydrophila. Immunoblotting experiments using the wild-type strain 569B and the secretion mutant M14 confirmed that ChiA is an extracellular protein which is secreted by the eps system. The eps system is also responsible for secreting cholera toxin, an oligomeric protein with no amino acid homology to ChiA, These results indicate that ChiA and cholera toxin have functionality similar extracellular transport signals that are essential for eps-dependent secretion.