Abstract
The DNA recombination and repair machineries of Mycoplasma genitalium and Mycoplasma pneumoniae differ considerably from those of gram-positive and gram-negative bacteria. Most notably, M. pneumoniae is unable to express a functional RecU Holliday junction (HJ) resolvase. In addition, the RuvB homologues from both M. pneumoniae and M. genitalium only exhibit DNA helicase activity but not HJ branch migration activity in vitro. To identify a putative role of the RuvA homologues of these mycoplasmas in DNA recombination, both proteins (RuvAMpn and RuvAMge, respectively) were studied for their ability to bind DNA and to interact with RuvB and RecU. In spite of a high level of sequence conservation between RuvAMpn and RuvAMge (68.8% identity), substantial differences were found between these proteins in their activities. First, RuvAMge was found to preferentially bind to HJs, whereas RuvAMpn displayed similar affinities for both HJs and single-stranded DNA. Second, while RuvAMpn is able to form two distinct complexes with HJs, RuvAMge only produced a single HJ complex. Third, RuvAMge stimulated the DNA helicase and ATPase activities of RuvBMge, whereas RuvAMpn did not augment RuvB activity. Finally, while both RuvAMge and RecUMge efficiently bind to HJs, they did not compete with each other for HJ binding, but formed stable complexes with HJs over a wide protein concentration range. This interaction, however, resulted in inhibition of the HJ resolution activity of RecUMge.
Highlights
A significant proportion of the genomes of Mycoplasma pneumoniae and Mycoplasma genitalium is composed of repeated DNA elements
These motifs were previously identified within domain II of RuvAEco and were shown to be crucial for sequence-independent DNA binding by interacting with the DNA phosphate backbone of Holliday junctions (HJs) [27,28,29]
As this RuvAMge-HJ complex had a similar mobility through polyacrylamide gels as RuvAMpn-HJ complex I and RuvAEco-HJ complex I [17,30,31,32,33,34,35], and because RuvAMge is a tetramer in solution, it is highly likely that this complex is composed of a tetramer of RuvAMge bound to a single HJ
Summary
A significant proportion of the genomes of Mycoplasma pneumoniae and Mycoplasma genitalium (approximately 8% and 4%, respectively) is composed of repeated DNA elements. These elements are referred to as RepMP elements in M. pneumoniae [1,2,3] and MgPa repeats (MgPars) in M. genitalium [4,5,6]. One or more of these variants are contained within open reading frames (ORFs) that encode antigenic surface proteins. Among these proteins are P1, P40 and P90 of M. pneumoniae and MgPa and P110 of M. genitalium. Homologous recombination between the repeated DNA elements in both Mollicutes species may play a crucial role in immune evasion [14]
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