Cross-complementation study of the flagellar type III export apparatus membrane protein FlhB.
Cross-complementation study of the flagellar type III export apparatus membrane protein FlhB.
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DOI：
10.1371/journal.pone.0044030
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发表时间：
2012
期刊：
影响因子：
3.7
通讯作者：
Samatey FA
中科院分区：
文献类型：
--
作者：
Barker CS;Samatey FA
III型细菌输出装置存在于一些致病性革兰氏阴性菌的鞭毛和针状复合体中。在针复合体中，其功能是分泌效应蛋白以感染真核细胞。在细菌鞭毛中，它输出特定的蛋白质，用于在组装过程中构建鞭毛。输出装置由大约五种膜蛋白和三种可溶性蛋白组成。出口装置的机制尚不完全清楚。这五种膜蛋白非常保守并且是必需的。这里进行了交叉互补测定：在沙门氏菌的鞭毛系统中，用来自 Aquifex aeolicus（一种进化遥远的超嗜热细菌）的 FlhB 直系同源物或由 A. aeolicus FlhB 的跨膜结构域与 沙门氏菌 FlhB 的细胞质结构域显着减少了鞭毛的数量和运动性。从表达嵌合 AquSalFlhB 蛋白的细胞中分离出运动性增强的抑制突变体，并使用全基因组测序鉴定突变。在编码 FlhA（III 型输出装置的另一种膜蛋白）的基因中发现了功能获得突变。此外，在编码 4-羟基苯甲酸辛基转移酶、泛醌/甲基萘醌生物合成甲基转移酶和 4-羟基-3-甲基丁-2-en-1-基二磷酸合酶（泛醌生物合成所需）的基因中也发现了突变。反相高效液相色谱显示突变减少了细胞质膜的醌池。通过添加过量的外源4-羟基苯甲酸酯，可以恢复带有4-羟基苯甲酸酯八异戊二烯基转移酶突变基因的菌株的泛醌生物合成。使用 AquSalFlhB 嵌合体恢复泛醌水平会减少鞭毛生物发生，表明呼吸链醌池是造成这种现象的原因。
The bacterial type III export apparatus is found in the flagellum and in the needle complex of some pathogenic Gram-negative bacteria. In the needle complex its function is to secrete effector proteins for infection into Eukaryotic cells. In the bacterial flagellum it exports specific proteins for the building of the flagellum during its assembly. The export apparatus is composed of about five membrane proteins and three soluble proteins. The mechanism of the export apparatus is not fully understood. The five membrane proteins are well conserved and essential. Here a cross-complementation assay was performed: substituting in the flagellar system of Salmonella one of these membrane proteins, FlhB, by the FlhB ortholog from Aquifex aeolicus (an evolutionary distant hyperthermophilic bacteria) or a chimeric protein (AquSalFlhB) made by the combination of the trans-membrane domain of A. aeolicus FlhB with the cytoplasmic domain of Salmonella FlhB dramatically reduced numbers of flagella and motility. From cells expressing the chimeric AquSalFlhB protein, suppressor mutants with enhanced motility were isolated and the mutations were identified using whole genome sequencing. Gain-of-function mutations were found in the gene encoding FlhA, another membrane protein of the type III export apparatus. Also, mutations were identified in genes encoding 4-hydroxybenzoate octaprenyltransferase, ubiquinone/menaquinone biosynthesis methyltransferase, and 4-hydroxy-3-methylbut-2-en-1-yl diphosphate synthase, which are required for ubiquinone biosynthesis. The mutations were shown by reversed-phase high performance liquid chromatography to reduce the quinone pool of the cytoplasmic membrane. Ubiquinone biosynthesis could be restored for the strain bearing a mutated gene for 4-hydroxybenzoate octaprenyltransferase by the addition of excess exogenous 4-hydroxybenzoate. Restoring the level of ubiquinone reduced flagella biogenesis with the AquSalFlhB chimera demonstrating that the respiratory chain quinone pool is responsible for this phenomenon.
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影响因子：
3.5
作者：
CHEREPANOV, PP;WACKERNAGEL, W
通讯作者：
WACKERNAGEL, W
影响因子：
14.9
作者：
Cole C;Barber JD;Barton GJ
通讯作者：
Barton GJ
DOI：
10.1073/pnas.92.6.1970
发表时间：
1995-03-14
影响因子：
11.1
作者：
GARZA, AG;HARRISHALLER, LW;MANSON, MD
通讯作者：
MANSON, MD
影响因子：
3.2
作者：
Kihara, M;Minamino, T;Macnab, RM
通讯作者：
Macnab, RM
影响因子：
2.8
作者：
Bekker, M.;Kramer, G.;de Mattos, M. J. Teixeira
通讯作者：
de Mattos, M. J. Teixeira