The bacterial conjugation protein TrwB resembles ring helicases and F1-ATPase

Abstract

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 plants or fungi and even bacterial sporulation 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 alpha/beta topology, reminiscent of RecA and DNA ring helicases, and an all-alpha domain. Six equivalent protein monomers associate to form an almost spherical quaternary structure that is strikingly similar to F1-ATPase. A central channel, 20 A in width, traverses the hexamer.

The mating pair formation system of conjugative plasmids—A versatile secretion machinery for transfer of proteins and DNA

“…Contrary to the dramatic rigidbody domain shifts between nucleotide-and unliganded forms of HP0525 (VirB11), nucleotidebound, and nucleotide-free TrwB N70 exhibits more subtle conformational changes (Gomis-Rüth et al, 2001. However, these structural rearrangements propagate all the way to the internal channel of the structure, suggesting that nucleotide-binding and/or hydrolysis may serve as a molecular switch facilitating the binding and threading of the protein-linked DNA substrate through the TrwB central channel during conjugation (Gomis-Rüth et al, 2001. The narrowing of the internal channel at the cytoplasmic face of the hexameric assembly however suggests that additional conformational adjustments must take place for widening this end of the channel, so that passage of the substrate can be achieved.…”

“…Similarly to the situation encountered with VirB4, the initially postulated NTPase activity of the CP has not been detected in vitro (Moncalián et al, 1999;Schröder et al, 2002). However, CPs have been demonstrated to bind nucleotides (NTPs) and also nucleotide diphosphates (NDPs) in vitro (Gomis-Rüth et al, 2001;Moncalián et al, 1999;Schröder and Lanka, 2003). Binding and release of nucleotides, which is possibly triggered by Mg 2+ , may have a mechanistic function related to substrate translocation (Schröder and Lanka, 2003).…”

“…The overall structure of TrwB N70 is remarkably similar to the structure of the F 1 -ATPase 3 3 heterohexamer (Abrahams et al, 1994) suggesting that T4CPs may also act as molecular motors. Contrary to the dramatic rigidbody domain shifts between nucleotide-and unliganded forms of HP0525 (VirB11), nucleotidebound, and nucleotide-free TrwB N70 exhibits more subtle conformational changes (Gomis-Rüth et al, 2001. However, these structural rearrangements propagate all the way to the internal channel of the structure, suggesting that nucleotide-binding and/or hydrolysis may serve as a molecular switch facilitating the binding and threading of the protein-linked DNA substrate through the TrwB central channel during conjugation (Gomis-Rüth et al, 2001.…”

“…As seen in crystal structures and electron microscopic images of the CP TrwB (plasmid R388), CPs form hexameric ring structures, reminiscent of ring helicases and F1-ATPase (Gomis-Rüth et al, 2001;Hormaeche et al, 2002). Other CPs (TraG of plasmid RP4, TraD of the F plasmid) form higher oligomers, when analyzed in vitro, but they may adopt a similar ring structure in vivo (Schröder et al, 2002).…”

Structure of a VirD4 coupling protein bound to a VirB type IV secretion machinery

“…VirB11 has been reported as a hexamer in different systems (Machon et al , 2002; Savvides et al , 2003; Hare et al , 2006). VirD4 and VirB4, though expected to form hexamers, have been identified in monomeric and dimeric forms as well (Gomis‐Ruth et al , 2001; Schroder et al , 2002; Rabel et al , 2003; Arechaga et al , 2008; Mihajlovic et al , 2009; Durand et al , 2011; Pena et al , 2012; Wallden et al , 2012). All three ATPases interact with one another and these interactions likely direct the two processes in which T4S systems are involved: pilus biogenesis and substrate secretion (Atmakuri et al , 2004; Ripoll‐Rozada et al , 2013).…”