Functional Characterization of the RuvB Homologs from Mycoplasma pneumoniae and Mycoplasma genitalium

Abstract

Homologous recombination between repeated DNA elements in the genomes of Mycoplasma species has been hypothesized to be a crucial causal factor in sequence variation of antigenic proteins at the bacterial surface. To investigate this notion, studies were initiated to identify and characterize the proteins that form part of the homologous DNA recombination machinery in Mycoplasma pneumoniae as well as Mycoplasma genitalium. A significant portion of the genomes of Mycoplasma pneumoniae and Mycoplasma genitalium consists of repeated DNA elements. This is remarkable as the genomes of these closely related human pathogens are relatively small, with lengths of 816 kb and 580 kb, respectively (6, 8). In M. pneumoniae, the repeated DNA elements are termed RepMP elements (29,37,44), whereas in M. genitalium, they are referred to as MgPa repeats (MgPars) (6,26,27). Although the RepMP elements and MgPa repeats do not have significant sequence homology, they do share two important features: (i) the different variants of these elements are similar in sequence but not identical, and (ii) one or more of these variants form part of open reading frames (ORFs) that code for antigenic surface proteins, such as the P1 protein in M. pneumoniae and the MgPa protein in M. genitalium. Because both P1 and MgPa can display sequence variation in the regions encoded by the RepMP or MgPar elements, it has been hypothesized that this variation is caused by homologous DNA recombination between the different variants of the RepMP or MgPar elements (10-11, 15, 17, 35, 36). Obviously, these recombination processes may supply a plethora of sequence variation to the P1 and MgPa genes, which can subsequently lead to amino acid sequence variation of the encoded antigenic proteins. Homologous DNA recombination between the repeated DNA elements in both Mycoplasma species may thus contribute significantly to evasion of these pathogens from the host's immune system.It is probable that the enzymatic machinery that governs recombination between repeated DNA elements in M. pneumoniae and M. genitalium largely overlaps with the machinery that is involved in homologous DNA recombination and DNA repair in these bacteria. Sequence analysis of the genomes of both M. pneumoniae and M. genitalium has revealed the presence of a distinct and limited set of ORFs that may encode the core proteins involved in DNA recombination pathways (2,6,8). The activities of some of these putative core proteins have already been investigated. These proteins include RuvA and a single-stranded DNA (ssDNA)-binding protein (SSB) from M. pneumoniae (9, 32) and the RecA and RecU proteins from both M. pneumoniae and M. genitalium (31,33,34). While the first three proteins exhibited activities that are similar to those of their counterparts from other bacteria, the RecU proteins from both Mycoplasma species displayed unusual and unique characteristics. First, while RecU from M. genitalium (RecU Mge ) was found to bind and cleave Holliday junction (HJ) substrates in a specific fashion, th...

Interaction of Branch Migration Translocases with the Holliday Junction-resolving Enzyme and Their Implications in Holliday Junction Resolution

“…In contrast, Mycoplasma genitalium RuvA bound to HJs, stimulated the DNA helicase and ATPase activities of RuvB, and did not compete RecU HJ binding (47). In the absence of RuvA, RuvB Mpn but not RuvB Mge showed helicase activity (55). It is likely, therefore, that although sharing similar proteins, the process of HJ resolution by the RuvAB-RecU machinery could act differently in the different bacteria and requires further analysis.…”

Characterization of the Operon Encoding the Holliday Junction Helicase RuvAB from Mycoplasma genitalium and Its Role in mgpB and mgpC Gene Variation

“…Orthologs of RuvA and RuvB in M. genitalium are encoded by genes MG_358 and MG_359 (referred to herein as ruvA and ruvB, respectively) (38,39). As with other bacterial species, both genes are adjacent and oriented in the same direction (Fig.…”

“…One example is the elevated helicase activity of the RuvB protein of the subtype 2 strain FH of M. pneumoniae (RuvB FH ) in the absence of RuvA (38). Similarly, RuvB of M. genitalium also displays intrinsic helicase activity, even though the activity levels are lower than that of RuvB FH , and this activity can be stimulated by RuvA (38,39).…”

“…Biochemical studies have shown that RuvA and RuvB proteins from M. genitalium and M. pneumoniae display in vitro activities that differ substantially from their counterparts in other bacteria (38,39). One example is the elevated helicase activity of the RuvB protein of the subtype 2 strain FH of M. pneumoniae (RuvB FH ) in the absence of RuvA (38).…”

“…One example is the elevated helicase activity of the RuvB protein of the subtype 2 strain FH of M. pneumoniae (RuvB FH ) in the absence of RuvA (38). Similarly, RuvB of M. genitalium also displays intrinsic helicase activity, even though the activity levels are lower than that of RuvB FH , and this activity can be stimulated by RuvA (38,39). Surprisingly, neither of these RuvB proteins can promote HJ branch migration in vitro irrespective of the presence after propagation of clone G37C, ⌬ruvA mutant, ⌬ruvB mutant, and complemented strains for 2, 4, 6, 8, and 10 consecutive passages (P2, P4, P6, P8, and P10).…”