Cell wall structure of a mutant of Mycobacterium smegmatis defective in the biosynthesis of mycolic acids.
Cell wall structure of a mutant of Mycobacterium smegmatis defective in the biosynthesis of mycolic acids.
复制标题
分枝杆菌酸生物合成缺陷的突变体的细胞壁结构。
DOI：
10.1074/jbc.275.10.7224
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发表时间：
2000
期刊：
影响因子：
--
通讯作者：
Liu,J
中科院分区：
文献类型：
--
作者：
Wang,L;Slayden,RA;Barry3rd,CE;Liu,J
最近分离出在分枝菌酸的生物合成中有缺陷的耻垢分枝杆菌的突变菌株（Liu，J.，和Nikaido，H.等人（1999）Proc. Acad. Sci.联合S. A.96，4011-4016）。该突变体不能合成全长分枝菌酸，并积累了一系列长链β-羟基分枝菌酸。在这项工作中，我们提供了一个详细的本地化的meromycolates和突变体的细胞壁结构的表征。薄层层析显示，用氯仿/甲醇和SDS提取后剩余的不溶性细胞壁基质仍然含有大部分的总meromycolates。基质辅助激光解吸/电离和电喷雾电离质谱分析所产生的史密斯降解的不溶性细胞壁基质的片段显示，meromycolates共价连接到阿拉伯半乳聚糖在5-OH位置的末端阿拉伯呋喃糖残基。通过NMR分析，阿拉伯半乳聚糖在突变株中似乎是正常的。有机相脂质的分析表明，突变体细胞壁含有一些可提取的脂质，但缺乏糖肽和脂寡糖。突变体细胞壁的差示扫描量热法未能显示完整的分枝杆菌细胞壁典型的大的合作热转变。透射电子显微镜观察发现，突变体细胞壁具有异常的超微结构（没有与不对称的真菌酸脂层相关的电子透明区）。综上所述，这些结果表明分枝菌酸对分枝杆菌细胞壁的结构和功能完整性的重要性。突变体细胞壁中缺乏高度组织化的脂质结构域解释了突变体的药物敏感性和温度敏感性表型。
A mutant strain ofMycobacterium smegmatisdefective in the biosynthesis of mycolic acids was recently isolated (Liu, J., and Nikaido, H. (1999)Proc. Natl. Acad. Sci. U. S. A.96, 4011–4016). This mutant failed to synthesize full-length mycolic acids and accumulated a series of long chain β-hydroxymeromycolates. In this work, we provide a detailed characterization of the localization of meromycolates and of the cell wall structure of the mutant. Thin layer chromatography showed that the insoluble cell wall matrix remaining after extraction with chloroform/methanol and SDS still contained a large portion of the total meromycolates. Matrix-assisted laser desorption/ionization and electrospray ionization mass spectroscopy analysis of fragments arising from Smith degradation of the insoluble cell wall matrix revealed that the meromycolates were covalently attached to arabinogalactan at the 5-OH positions of the terminal arabinofuranosyl residues. The arabinogalactan appeared to be normal in the mutant strain, as analyzed by NMR. Analysis of organic phase lipids showed that the mutant cell wall contained some of the extractable lipids but lacked glycopeptidolipids and lipooligosaccharides. Differential scanning calorimetry of the mutant cell wall failed to show the large cooperative thermal transitions typical of intact mycobacterial cell walls. Transmission electron microscopy showed that the mutant cell wall had an abnormal ultrastructure (without the electron-transparent zone associated with the asymmetric mycolate lipid layer). Taken together, these results demonstrate the importance of mycolic acids for the structural and functional integrity of the mycobacterial cell wall. The lack of highly organized lipid domains in the mutant cell wall explains the drug-sensitive and temperature-sensitive phenotypes of the mutant.