Cellular localization and functional analysis of the protein encoded by the chromosomal virulence gene(acvB) of Agrobacterium tumefaciens

Abstract

A chromosomal virulence gene, acvB, of Agrobacterium tumefaciens [J. Bacteriol., 175, 3208-3212 (1993)] was over-expressed in Escherichia coli. A 47-kDa protein was produced and localized in the periplasmic space of E. coli. Amino acid sequence analysis of its N-terminal demonstrated that a signal peptide of 24 amino acids was cleaved from the pre AcvB protein to produce the mature 47-kDa protein. Western-blot analysis using the antiserum against the AcvB protein detected a 47-kDa protein in the periplasmic space only with strain A208 (acvB+). The amount of AcvB protein synthesized was not increased in strain A208 by induction with acetosyringone (100 microM). There was observed no significant difference in induction by acetosyringone of virB::lacZ, virD::lacZ, and virE::lacZ fusion genes regardless of the presence or absence of the acvB gene. The T-strand (lower strand of T-DNA) was detected in strains A208 as well as B119 (acvB-) which were cultured in induction medium containing acetosyringone. AcvB protein bound to single-stranded DNAs with no apparent sequence specificity. The results suggest that AcvB protein binds to the T-strand in periplasm and mediates the transfer of the T-strand from A. tumefaciens to the host plant cell.