R388 cosmid vector广宿主接合转移载体

The transfer of DNA across membranes and between cells is a central biological process; however, its molecular mechanism remains unknown. In prokaryotes, trans-membrane passage by bacterial conjugation, is the main route for horizontal gene transfer. It is the means for rapid acquisition of new genetic information, including antibiotic resistance by pathogens. Trans-kingdom gene transfer from bacteria to plants¹ or fungi² and even bacterial sporulation³ are special cases of conjugation. An integral membrane DNA-binding protein, called TrwB in the Escherichia coli R388 conjugative system, is essential for the conjugation process. This large multimeric protein is responsible for recruiting the relaxosome DNA–protein complex, and participates in the transfer of a single DNA strand during cell mating. Here we report the three-dimensional structure of a soluble variant of TrwB. The molecule consists of two domains: a nucleotide-binding domain of / topology, reminiscent of RecA and DNA ring helicases, and an all- domain. Six equivalent protein monomers associate to form an almost spherical quaternary structure that is strikingly similar to F₁-ATPase. A central channel, 20 Å in width, traverses the hexamer.

R388 is a 33-kilobase enterobacterial conjugative plasmid that contains genes that code for sulphonamide and trimethoprim resistance.

Map:

A: genetic organization of plasmid R388. ORFs and other sequence features are depicted in different colors according to the legend on the left, after assignment to functional modules based on database similarity and/or available experimental data [12]. The R388 genome is compact and organized in two major modules, one devoted to general maintenance functions and one devoted to conjugation (MOB, Mobility genes; MPF, Mating Pore Formation). stbDRs, located around the promoter region of the stbABC operon and consisting of two sets of five direct repeats of a 9 bp DNA sequence spaced by two nucleotides, are indicated. This 9 bp DNA sequence is also present in number of two or three in three other promoters of the maintenance region. B: Synteny conservation between the MOB-Stb region of R388 and those of other plasmids. Hatched arrows indicate the relaxase gene. Plasmid GenBank accession numbers: R388: BR000038, R46: NC_003292, NAH7: NC_007926, RP4: L27758, pTF-FC2: M57717.1, R721: NC_002525.