The lipoprotein VirB7 interacts with VirB9 in the membranes of Agrobacterium tumefaciens
The lipoprotein VirB7 interacts with VirB9 in the membranes of Agrobacterium tumefaciens
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DOI：
10.1128/jb.179.4.1211-1218.1997
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发表时间：
1997-02-01
影响因子：
3.2
通讯作者：
Zambryski, PC
中科院分区：
文献类型：
--
作者：
Baron, C;Thorstenson, YR;Zambryski, PC
VirB9和VirB7是推测的将t复合物从农杆菌转移到植物中所需的VirB膜通道的重要组成部分。在这篇报告中，我们提出了它们相互作用和细胞定位的生化分析。在非还原和还原条件下的相对电泳迁移率的比较表明，它们与其他蛋白质形成巯基敏感复合物。双向凝胶电泳鉴定出一个复合体为VirB9和VirB7二硫键共价连接的异源二聚体，以及VirB7同型二聚体和单体。virb9特异性抗血清免疫沉淀分离出异二聚体VirB9-VirB7复合物。用还原剂孵育将复合体分裂为VirB9和VirB7，进一步证实了通过半胱氨酸残基连接。在酵母双杂交系统中也观察到VirB9和VirB7的相互作用。VirB9的膜附着可能是由于脂蛋白修饰，因为在a . tummefaciens中用[H-3]棕榈酸标记证实了VirB7是与VirB9相关的脂蛋白。VirB9和VirB7在内膜和外膜之间分布均匀，这与它们与跨膜VirB复合物的关联一致。
VirB9 and VirB7 are essential components of the putative VirB membrane channel required for transfer of the T-complex from Agrobacterium tumefaciens into plants. In this report, we present a biochemical analysis of their interaction and cellular localization. A comparison of relative electrophoretic mobilities under nonreducing and reducing conditions suggested that they form thiol-sensitive complexes with other proteins. Two-dimensional gel electrophoresis identified one complex as a heterodimer of VirB9 and VirB7 covalently linked by a disulfide bond, as well as VirB7 homodimers and monomers. Immunoprecipitation with VirB9-specific antiserum isolated the heterodimeric VirB9-VirB7 complex. Incubation with reducing agent split the complex into its constituent VirB9 and VirB7, which further confirmed linkage via cysteine residues. The interaction between VirB9 and VirB7 also was observed in the yeast two-hybrid system. Membrane attachment of VirB9 VirB7 may be conferred by lipoprotein modification, since labeling with [H-3]palmitic acid in A. tumefaciens verified that VirB7 is a lipoprotein associated with VirB9. VirB9 and VirB7 showed equal distribution between inner and outer membranes, in accord with their proposed association with the transmembrane VirB complex.