Fig. 4
Site-directed mutagenesis on TraE and localization on crystal structure. (A) Amino acids sequence alignment of TraE from pKM101 plasmid with six VirB8 homologs: B. suis (strain 1330, 38% identity), A. tumefaciens (C58 plasmid, 27% identity), E. coli R388 plasmid (42% identity), S. meliloti (SM11 plasmid, 26% identity), B. henselae (ICB.4D7 plasmid, 21% identity), and A. baumannii (Rp428, 35% identity). Strongly positively charged and aromatic amino acids residues are marked in grey. (B) pKM101 conjugation assay to assess the capacity of TraE and its variants to complement a pKM101ΔtraE plasmid. The data represent averages and standard errors of three biological replicate cultures. Asterisks represent statistically significant differences (**** for p < 0.0001, n = 3; *** for p < 0.001, n = 3; ** for p < 0.01, n = 3; ns for non significant, n = 3). Bottom, Western blot using anti-TraE antibody. (C) Crystal structure of TraE dimer (pdb: 5I97) with location of mutation sites. Molecular graphics and analyses were performed with UCSF Chimera (https://www.rbvi.ucsf.edu/chimera), developed by the Resource for Biocomputing, Visualization, and Informatics at the University of California., San Francisco, with support from NIH P41-GM103311.
