THE LIPOPROTEIN VIRB7 INTERACTS WITH VIRB9 IN THE MEMBRANES OF AGROBACTERIUM-TUMEFACIENS

VirB9 and VirB7 are essential components of the putative VirB membrane channel required for transfer of the T-complex from Agrobacterium tum efaciens into plants. In this report, we present a biochemical analysi s of their interaction and cellular localization. A comparison of rela tive electrophoretic mobilities under nonreducing and reducing conditi ons suggested that they form thiol-sensitive complexes with other prot eins. 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 Vir B9-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 yea st two-hybrid system. Membrane attachment of VirB9 VirB7 may be confer red by lipoprotein modification, since labeling with [H-3]palmitic aci d in A. tumefaciens verified that VirB7 is a lipoprotein associated wi th VirB9. VirB9 and VirB7 showed equal distribution between inner and outer membranes, in accord with their proposed association with the tr ansmembrane VirB complex.