PDBsum entry 3ewa

DNA binding protein, recombination

PDB id: 3ewa

Contents

Protein chain

314 a.a.

Ligands

ANP

Metals

_MG ×2

Waters ×45

References listed in PDB file

Key reference

Title

Conservation of a conformational switch in rada recombinase from methanococcus maripaludis.

Authors

Y.Li, Y.He, Y.Luo.

Ref.

Acta Crystallogr D Biol Crystallogr, 2009, 65, 602-610. [DOI no: 10.1107/S0907444909011871]

PubMed id

19465774

Abstract

Archaeal RadAs are close homologues of eukaryal Rad51s ( approximately 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'-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 DNA-interacting 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.

Figure 4.
Figure 4 Superimposed ATPase centres of MmRadA and EcRecA in stereo. The conserved P-loops are superimposed. The P-loop (Gly105-Thr112), His280 and Asp302 of the ATPase-active form of MmRadA are shown as green stick models. The MmRadA-bound AMPPNP and monovalent cations are shown as yellow ball-and-stick models. The P-loop (Gly66-Thr73), Phe217, Lys248 and Lys250 of the active EcRecA structure are shown in blue.

Figure 5.
Figure 5 Hypothetical model of ATP hydrolysis-facilitated gap displacement. The crystallized protein filament is shown as a C^ trace in salmon with AMPPNP in green. Speculative models of the DNA substrates are shown as wires. Important segments are boxed. The 5'- and 3'-ends are based on the filament-initiating ssDNA (thinner wire in blue). The homologous dsDNA is shown as a thicker wire in yellow. The strand-exchange process progresses from the 5'-end to the 3'-end. (a) An intervening gap. Such gaps are likely to exist owing to simultaneous homologous pairing between the recombinase/ssDNA filament and dsDNA at multiple locations. The dsDNA in the gap region cannot become properly wound ( 19 bp per helical turn) around the nucleoprotein filament without unwinding its adjacent region(s). Despite the sequence homology, it serves as a topological roadblock of strand exchange between long DNA substrates. (b) ATP hydrolysis promotes the transient release of a dsDNA segment at the immediate 3'-flank of the gap. The transiently released dsDNA region is shown as an exaggerated wide helix. (c) Rearrangement in the transiently released dsDNA region and the adjacent gap takes place without changing the overall topology. The 5'-end of the gap region becomes properly wound, while the released dsDNA region becomes unwound. (d) The rearranged 5'-end of the gap becomes bound by the recombinase filament. As a result, the gap is displaced towards the 3'-end. (e) Repetition of steps (b)-(d) would chase the topologically strained gap out of the 3'-end of the nucleoprotein filament, therefore removing topological roadblocks to extensive DNA strand exchange.

The above figures are reprinted from an Open Access publication published by the IUCr: Acta Crystallogr D Biol Crystallogr (2009, 65, 602-610) copyright 2009.