The Agrobacterium tumefaciens virB7 gene product, a proposed component of the T-complex transport apparatus, is a membrane-associated lipoprotein exposed at the periplasmic surface
The Agrobacterium tumefaciens virB7 gene product, a proposed component of the T-complex transport apparatus, is a membrane-associated lipoprotein exposed at the periplasmic surface
复制标题
DOI：
10.1128/jb.178.11.3156-3167.1996
复制
发表时间：
1996-06-01
影响因子：
3.2
通讯作者：
Christie, PJ
中科院分区：
文献类型：
--
作者：
Fernandez, D;Dang, TAT;Christie, PJ
根癌土壤杆菌virB 7基因产物含有以细菌脂蛋白特征性的共有信号肽酶II切割位点结束的典型信号序列。通过（i）体内标记天然VirB 7和VirB 7：：PhoA与[H-3]棕榈酸的融合和（ii）球霉素对VirB 7加工的抑制，球霉素是一种已知的信号肽酶II的抑制剂，一种VirB 7衍生物，在信号肽酶II切割位点内维持Ser取代不变的Cys-15残基，在免疫学上不能被观察到，并且不能补充Delta virB 7突变，从而确立了这种推定的脂质附着位点对VirB 7成熟和功能的重要性，VirB 7主要用来自野生型A348细胞的外膜级分以及用virB 7表达质粒转化的Delta virB操纵子衍生物分配，与phoA融合的virB 7的表达，Escherichia coli碱性磷酸酶基因，在E. coli和A.根癌农杆菌细胞，为VirB 7在这些宿主中的输出提供遗传证据，VirB 7被证明对蛋白酶K具有内在抗性;相反，VirB 7：：PhoA衍生物被A的蛋白酶K处理降解。总之，这些研究的结果支持这样的模型，其中VirB 7在拓扑学上被配置为单聚蛋白，其氨基末端主要锚定在外膜上，其亲水性羧基结构域位于周质空间中。VirB 10证实这些膜相关蛋白中的每一种也含有大的周质结构域，而VirB 11主要或专门存在于细胞内部。
The Agrobacterium tumefaciens virB7 gene product contains a typical signal sequence ending with a consensus signal peptidase II cleavage site characteristic of bacterial lipoproteins. VirB7 was shown to he processed as a lipoprotein by (i) in vivo labeling of native VirB7 and a VirB7::PhoA fusion with [H-3] palmitic acid and (ii) inhibition of VirB7 processing by globomycin, a known inhibitor of signal peptidase II, A VirB7 derivative sustaining a Ser substitution for the invariant Cys-15 residue within the signal peptidase II cleavage site could not be visualized immunologically and failed to complement a Delta virB7 mutation, establishing the importance of this putative lipid attachment site for VirB7 maturation and function, VirB7 partitioned predominantly with outer membrane fractions from wild-type A348 cells as well as a Delta virB operon derivative transformed with a virB7 expression plasmid, Expression of virB7 fused to phoA, the alkaline phosphatase gene of Escherichia coli, gave rise to high alkaline phosphatase activities in E. coli and A. tumefaciens cells, providing genetic evidence for the export of VirB7 in these hosts, VirB7 was shown to be intrinsically resistant to proteinase K; by contrast, a VirB7::PhoA derivative was degraded by proteinase K treatment of A. tumefaciens spheroplasts and remained intact upon treatment of whole cells, Together, the results of these studies favor a model in which VirB7 is topologically configured as a monotopic protein with its amino terminus anchored predominantly to the outer membrane and with its hydrophilic carboxyl domain located in the periplasmic space, Parallel studies of VirB5, VirB8, VirB9, and VirB10 established that each of these membrane-associated proteins also contains a large periplasmic domain whereas VirB11 resides predominantly or exclusively within the interior of the cell.