Structure of a hexameric form of RadA recombinase fromMethanococcus voltae

Abstract

PDB Reference: RadA recombinase, 4dc9.Archaeal RadA proteins are close homologues of eukaryal Rad51 and DMC1 proteins and are remote homologues of bacterial RecA proteins. For the repair of double-stranded breaks in DNA, these recombinases promote a pivotal strand-exchange reaction between homologous single-stranded and doublestranded DNA substrates. This DNA-repair function also plays a key role in the resistance of cancer cells to chemotherapy and radiotherapy and in the resistance of bacterial cells to antibiotics. A hexameric form of a truncated Methanococcus voltae RadA protein devoid of its small N-terminal domain has been crystallized. The RadA hexamers further assemble into two-ringed assemblies. Similar assemblies can be observed in the crystals of Pyrococcus furiosus RadA and Homo sapiens DMC1. In all of these two-ringed assemblies the DNA-interacting L1 region of each protomer points inward towards the centre, creating a highly positively charged locus. The electrostatic characteristics of the central channels can be utilized in the design of novel recombinase inhibitors.

Structure of a filament of stacked octamers of human DMC1 recombinase

“…In the structures of filamentous or hexameric RadA proteins from M. voltae and M. maripaludis, disorder of helix G has been correlated with either the presence of post-hydrolysis ADP or the absence of proper cationic bridging between the C-terminal carbonyl groups of helix G and the -phosphate of the ATP analog AMPPNP. This structural feature of Á 83 HsDMC1 is consistent with the notions that helix G is intrinsically stable (Du & Luo, 2012) and that ATP hydrolysis in recombinase filaments triggers disordering of helix G and further renders the L2 region incompatible for binding DNA (Qian et al, 2005;Li et al, 2009).…”

“…Crystallized 'inactive' EcRecA filaments with shorter helical pitches were also observed two decades ago (Story et al, 1992). Ring-shaped 'inactive' forms of such recombinases with 6-8 protomers have also commonly been observed by electron microscopy (Heuser & Griffith, 1989;Yu & Egelman, 1997;Passy et al, 1999;Yang et al, 2001;Galkin et al, 2006;McIlwraith et al, 2001;Masson et al, 1999) and by crystallography (Shin et al, 2003;Kinebuchi et al, 2004;Du & Luo, 2012). In addition to the three commonly found forms, crystal structures of overwound three-monomer-per-turn filaments (Ariza et al, 2005) and lefthanded filaments of Sulfolobus solfataricus RadA (Chen et al, 2007;Chang et al, 2009) have also been observed.…”

“…Such domains start with a polymerization motif (Pellegrini et al, 2002) centered around a hydrophobic residue (Phe85 in HsDMC1) which protrudes into a hydrophobic pocket in an adjacent monomer in the recombinase polymer. Similarly truncated ATPase domains of RadA from Methanococcus voltae (MvRadA) have been crystallized as either helical filaments (Galkin et al, 2006) or hexamers (Du & Luo, 2012). As observed in full-length HsDMC1, the Á 83 HsDMC1 protein also formed octamers in the crystal.…”

Pan-Genome Analysis of Human Gastric PathogenH. pylori: Comparative Genomics and Pathogenomics Approaches to Identify Regions Associated with Pathogenicity and Prediction of Potential Core Therapeutic Targets

“…However, some genes on these PAIs deserve attention; for example, PiHp2 harbors a gene coding for a DNA repair protein, RadA (C694_01125), which is part of a superfamily of recombinases or DNA strand-exchange proteins, composed of archaeal RadA, bacterial RecA, and eukaryal Rad51 and DMC1 proteins. RadA has a pivotal role in DNA strand-exchange process between single stranded DNA (ssDNA) and a homologous double-stranded DNA (ds-DNA) in homologous recombination [ 47 ]. Homologous recombination is one of several DNA repair pathways (direct reversal, base excision repair, nucleotide excision repair, mismatch repair, and recombination repair pathways) and functions in the repair of double-stranded DNA breaks and the restarting of stalled replication forks, therefore, guaranteeing accurate functioning and propagation of genetic information [ 47 , 48 ].…”

“…RadA has a pivotal role in DNA strand-exchange process between single stranded DNA (ssDNA) and a homologous double-stranded DNA (ds-DNA) in homologous recombination [ 47 ]. Homologous recombination is one of several DNA repair pathways (direct reversal, base excision repair, nucleotide excision repair, mismatch repair, and recombination repair pathways) and functions in the repair of double-stranded DNA breaks and the restarting of stalled replication forks, therefore, guaranteeing accurate functioning and propagation of genetic information [ 47 , 48 ]. However, as RadA is mainly found in archaea, in vitro experiments are required to elucidate the putative function of RadA in bacteria, specially H. pylori and to clarify its putative acquirement through horizontal gene transfer from archaea.…”