Conservation of a conformational switch in RadA recombinase fromMethanococcus maripaludis

Abstract

Archaeal RadAs are close homologues of eukaryal Rad51s ($40% sequence identity). These recombinases promote ATP hydrolysis and a hallmark strand-exchange reaction between homologous single-stranded and double-stranded DNA substrates. Pairing of the 3 0 -overhangs located at the damaged DNA with a homologous double-stranded DNA enables the re-synthesis of the damaged region using the homologous DNA as the template. In recent studies, conformational changes in the DNA-interacting regions of Methanococcus voltae RadA have been correlated with the presence of activity-stimulating potassium or calcium ions in the ATPase centre. The series of crystal structures of M. maripaludis RadA presented here further suggest the conservation of an allosteric switch in the ATPase centre which controls the conformational status of DNA-interacting loops. Structural comparison with the distant Escherichia coli RecA homologue supports the notion that the conserved Lys248 and Lys250 residues in RecA play a role similar to that of cations in RadA. The conservation of a cationic bridge between the DNAinteracting L2 region and the terminal phosphate of ATP, together with the apparent stability of the nucleoprotein filament, suggests a gap-displacement model which may explain the advantage of ATP hydrolysis for DNA-strand exchange.

Ancient systems of sodium/potassium homeostasis as predecessors of membrane bioenergetics

“…The bacterial homolog of RadA is the RecA recombinase. A comparison of the crystal structures of RadA from Methanococcus maripaludis (PDB ID 3EW9) and E. coli RecA [55] shows that, after superposition of the P-loops, the K + ions bound by RadA overlay the lysine residues K248 and K250 of RecA, which are important for catalysis [56]. Although these residues are located on the surface of the RecA monomer that is far away from the ATP binding site, they are inserted into the ATP binding site of the next monomer in the 3D structure of the complex with DNA.…”

“…The α-subunit of bovine rotary ATP synthase (PDB 1COW [153], shown in gray) and the RecA protein from E. coli (PDB 3CMX [55], shown in light violet) were superimposed with the PDBeFold software [154]. The superimposed part is shown in dark violet.…”

Structure of a hexameric form of RadA recombinase fromMethanococcus voltae

“…These proteins facilitate a pivotal DNA strand-exchange process between a single-stranded DNA (ssDNA) and a homologous double-stranded DNA (dsDNA) in homologous recombination. Electron-microscopic and crystallographic results have revealed strikingly similar 'active' recombinase assemblies in the form of right-handed helical filaments with approximately six monomers per turn (VanLoock et al, 2003;Conway et al, 2004;Wu et al, 2004;Chen et al, 2008;Sheridan et al, 2008;Li et al, 2009a). The milestone structures of Escherichia coli RecA (EcRecA) in complex with a series of DNA molecules have shed light on the exact mechanism of homologous DNA strand exchange (Chen et al, 2008).…”

RAD51 Protein ATP Cap Regulates Nucleoprotein Filament Stability

“…6D) (43). The structure of MvRAD51(D302K) closely resembles the recurrent active form of M. voltae and Methanococcus maripaludis RAD51⅐AMP-PNP complexes (18,44,45) but differs from the recurrent inactive form of MvRAD51 (17), which has a largely disordered L2 region (compare Fig. 6C with Fig.…”