Characterization of trbC, a new F plasmid tra operon gene that is essential to conjugative transfer

Abstract

We have characterized a previously unidentified gene, trbC, which is contained in the transfer region of the Escherichia coi K-12 fertility factor, F. Our data show that the trbC gene is located between the F plasmid genes traU and traN. The product of trbC was identified as a polypeptide with an apparent molecular weight (M. amino-terminal signal sequence. We also constructed two different Km' gene insertion mutations in trbC and crossed these onto the transmissible F plasmid derivative pOX38. We found that cells carrying pOX38 trbC mutant plasmids were transfer deficient and resistant to infection by F-pilus-specific phages. Transfer proficiency and bacteriophage sensitivity were restored by complementation when a trbC+ plasmid clone was introduced into these cells. These results showed that trbC function is essential to the F plasmid conj,ugative

Characterization, localization, and sequence of F transfer region products: the pilus assembly gene product TraW and a new product, TrbI

“…Vectors used were * Corresponding author. pACYC177 (4), pLa2311 (21), and pKI497 (15). The latter is a derivative of pTZ18U (16) which carries tcy in place of lacZot.…”

“…The Tn9O3 kanamycin resistance (kan) gene cassette used came from plasmid pUC4K (Pharmacia Inc., Molecular Biology Division, Piscataway, N.J.). Matings and selections used for in vivo construction of pOX38 mutant derivatives were as previously described (11,15). pOX38 is a transmissible F replicon made by recircularization of the large F HindIII fragment and lacks transposable sequences (6).…”

“…Plasmid products were labeled with [35S]methionine in SE5000 maxicells, separated on sodium dodecyl sulfatepolyacrylamide gradient gels, and identified by autoradiography. When noted, the maxicell samples were fractionated prior to analysis on the gels to obtain periplasmic proteins (15) or inner membranes (13).…”

“…F lac and F lac traW546 strains have been described elsewhere (14). Host (13,15). Plasmid products were labeled with [35S]methionine in SE5000 maxicells, separated on sodium dodecyl sulfatepolyacrylamide gradient gels, and identified by autoradiography.…”

The identification of exported proteins with gene fusions to invasin

“…It is interesting that SrgA is an outer membrane-associated lipoprotein (Table 2), whereas E. coli DsbA is free within the periplasm (Bardwell et al, 1991). Homologues of two additional periplasmic proteins involved in pilus assembly, TrbC and TraW (Maneewannakul et al, 1991;Neidhardt, 1996) Error bars represent the standard deviation. After three cycles of selection, each isolate analysed was three logs more invasive that the background strain containing pICOM II without an insert.…”