Sturm S, Jann B, Jann K, Fortnagel P, Timmis K N
Department of Medical Biochemistry, University of Geneva, Switzerland.
Microb Pathog. 1986 Jun;1(3):307-24. doi: 10.1016/0882-4010(86)90056-2.
The genetic organization and functions of the Shigella dysenteriae 1 rfb gene cluster, which specifies the somatic O antigen in this organism, have been studied in Escherichia coli K-12 by insertion and deletion mutagenesis of pSS9, a pBR322 hybrid containing the Shigella rfb genes. On the basis of the sensitivity/resistance to rough-specific bacteriophage T3 of E. coli K-12 derivatives containing mutant pSS9 plasmids, of the banding patterns and immunoreactivity of LPS isolated from such derivatives and electrophoresed on SDS-polyacrylamide gels, and of the sugar composition of the polysaccharide portion of the LPS determined by chemical analysis, six determinants for O antigen production were identified and localized. At least two determinants are involved in synthesis of TDP-rhamnose and the transfer of a rhamnose residue to the galactose-substituted core. One of these functions is probably TDP-rhamnose synthetase. A third function effects the transfer of a second rhamnose residue to the rha----gal-substituted core. A fourth function, for which evidence was obtained for two determinants (cistrons), is N-acetylglucosamine transferase, whereas a sixth determinant is necessary for extension of the first completed side chain repeat unit to the full O antigen polymer. These results confirmed the previously-determined chemical composition of the S. dysenteriae 1 O antigen and demonstrated that the order of the sugars is glcNAc----rha----rha----gal with gal as the first sugar linked to the core. Evidence was obtained for at least two transcriptional units in the rfb gene cluster and the approximate locations of two promoters are suggested. The detection of new electrophoretic species of LPS that may correspond to LPS biosynthetic intermediates, and the finding on the cell surfaces of structures corresponding to LPS core substituted with one or more O-specific sugars, appear to be novel findings.
痢疾志贺氏菌1型rfb基因簇决定了该菌的体细胞O抗原，其基因组织和功能已通过对pSS9进行插入和缺失诱变在大肠杆菌K-12中进行了研究，pSS9是一种含有志贺氏菌rfb基因的pBR322杂种质粒。基于含有突变pSS9质粒的大肠杆菌K-12衍生物对粗糙特异性噬菌体T3的敏感性/抗性、从此类衍生物中分离并在SDS-聚丙烯酰胺凝胶上电泳的脂多糖的条带模式和免疫反应性，以及通过化学分析确定的脂多糖多糖部分的糖组成，鉴定并定位了六个O抗原产生的决定因素。至少有两个决定因素参与TDP-鼠李糖的合成以及鼠李糖残基向半乳糖取代核心的转移。其中一个功能可能是TDP-鼠李糖合成酶。第三个功能影响第二个鼠李糖残基向鼠李糖-半乳糖取代核心的转移。第四个功能（已获得两个决定因素（顺反子）的证据）是N-乙酰葡糖胺转移酶，而第六个决定因素对于将第一个完整的侧链重复单元延伸至完整的O抗原聚合物是必需的。这些结果证实了先前确定的痢疾志贺氏菌1型O抗原的化学组成，并表明糖的顺序是GlcNAc-鼠李糖-鼠李糖-半乳糖，半乳糖是与核心相连的第一个糖。在rfb基因簇中获得了至少两个转录单元的证据，并推测了两个启动子的大致位置。检测到可能对应于脂多糖生物合成中间体的新的脂多糖电泳种类，以及在细胞表面发现对应于被一个或多个O特异性糖取代的脂多糖核心的结构，似乎是新的发现。
