Chitin catabolism in the marine bacterium Vibrio furnissii -: Identification and molecular cloning of a chitoporin
Chitin catabolism in the marine bacterium Vibrio furnissii -: Identification and molecular cloning of a chitoporin
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DOI：
10.1074/jbc.m001041200
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发表时间：
2000-10-20
影响因子：
4.8
通讯作者：
Roseman, S
中科院分区：
文献类型：
--
作者：
Keyhani, NO;Li, XB;Roseman, S
由海洋细菌Vibrioelissii催化的几丁质涉及许多基因和蛋白质，包括两种独特的周质水解酶，壳聚糖糊精酶和β-N-乙酰氨基葡糖苷酶（Keyhani，N. O.，和Roseman，S，（1996）J. Biol; Chem.271，33414-33424和33425-33432）。外膜中的特异性壳孔蛋白可能是这些糖苷酶接近由几丁质酶产生的细胞外壳寡糖（GlcN-Ac）（n）所必需的。我们在这里报告的鉴定和分子克隆这样的孔蛋白。当葡萄糖胺酸（GlcNAc）（n = 2-6）诱导时，外膜蛋白OMP（表观分子量40 kDa）表达，但不被GlcNAc或其它糖诱导。根据OMP的N端序列，合成寡核苷酸并用于克隆基因chiP。Chip的推导氨基酸序列与几种细菌孔蛋白相似; OMP是Chip的加工形式。在大肠杆菌中，观察到两种重组蛋白，对应于加工和未加工的形式的ChiP。与亲本菌株相反，突变体不生长在（GlcNAc）上，并以降低的速率运输（GlcNAc）的非代谢类似物。这些结果表明，芯片是一个特定的chitoporin。
Chitin catabolism by the marine bacterium Vibrio furnissii involves many genes and proteins, including two unique periplasmic hydrolases, a chitodextrinase and a beta -N-acetylglucosaminidase (Keyhani, N. O., and Roseman, S, (1996) J. Biol; Chem. 271, 33414-33424 and 33425-33432). A specific chitoporin in the outer membrane may be required for these glycosidases to be accessible to extracellular chitooligosaccharides, (GlcN-Ac)(n), that are produced by chitinases. We report here the identification and molecular cloning of such a porin. An outer membrane protein, OMP (apparent molecular mass 40 kDa) was expressed when V, furnissii was induced by (GlcNAc)(n), n = 2-6, but not by GlcNAc or other sugars. Based on the N-terminal sequence of OMP, oligonucleotides were synthesized and used to clone the gene, chiP. The deduced amino acid sequence of ChiP is similar to several bacterial porins; OMP is a processed form of Chip. In Escherichia coli, two recombinant proteins were observed, corresponding to processed and unprocessed forms of ChiP, A null mutant of chiP was constructed in V. furnissii. In contrast to the parental strain, the mutant did not grow on (GlcNAc), and transported a nonmetabolizable analogue of (GlcNAc), at a reduced rate. These results imply that ChiP is a specific chitoporin.