RecA Protein

Abstract

The RecA protein of Escherichia coli is the prototypic deoxyribonucleic acid (DNA)‐strand exchange protein. It assembles on single‐stranded DNA to form a helical nucleoprotein filament that is the active species for all RecA protein‐dependent functions. This protein–DNA complex is responsible for three mutually exclusive functions: DNA recombination, induction of the DNA‐damage SOS response and SOS‐induced mutagenesis. The overall structure and activity of the nucleoprotein filament are determined by nucleoside triphosphate (NTP) cofactor binding, hydrolysis and product release. This is evident when the crystal structure of the filament is closely examined and NTP‐related structural changes are related to biological function.

Key Concepts

  • RecA protein is the prototype enzyme for a family of proteins known as recombinases.
  • Homologous recombination is the exchange of the corresponding stretches of DNA between two DNA molecules of similar sequence.
  • The SOS response is an inducible DNA repair and damage tolerance system that is induced by RecA.
  • SOS‐induced mutagenesis is a form of DNA repair catalysed by error‐prone, translesion DNA polymerases.
  • The RecA nucleoprotein filament is a large dynamic structure whose formation is essential to all RecA functions.
  • Coprotease activity is the induced self‐cleavage of one protein (LexA) by another (RecA).

Keywords: DNA‐strand exchange; genetic recombination; SOS response; SOS mutagenesis; nucleoprotein filament; LexA repressor

Figure 1. Roles of the RecA nucleoprotein filament in DNA metabolism. A schematic showing the three mutually exclusive functions of the RecA protein filament. The LexA–DNA complex is from Zhang et al. and the docked RecA–LexA complex is taken from Butala et al. . M. Butala (University of Ljubljana, Slovenia) provided the coordinates for the docked complex. The structure of the RecA nucleoprotein filament and the three‐strand product complex in homologous recombination are adapted from Chen et al. . Coordinates for the UmuC model were provided by M. Goodman (University of Southern California, USA) and were published in Gruber et al. and those of UmuD′ are from Peat et al. . Structures were artistically docked to create Pol V and the mutasomes.
Figure 2. ATP binding converts RecA from the inactive to the active state. (a) A single turn of the filament is shown in schematic form. Inset: the location of the ATP binding site at the monomer–monomer interface. The 5′‐proximal RecA is coloured in cyan, with the Walker A and B residues coloured yellow and pale pink, respectively. K72, which is the key residue in the Walker A box, is highlighted in green for clarity. When this reside us mutated, ATPase activity is essentially abolished. Red, ATP. The 3′‐distal monomer is coloured orange and key lysine residues from this subunit involved in ATPase activity are coloured purple. Coordinates for this image are from Chen et al. . (b,c) Three‐dimensional reconstructions of electron micrographs of RecA filaments formed on dsDNA with ADP (b) or ATPγS (c) (VanLoock et al., ). (d,e) Single turns of the RecA filament as seen in the crystal structure (Chen et al., ), (d) Inactive and (e) active filaments. In the absence of ATP or in the presence of ADP, the RecA filament is in a collapsed or inactive state (left side of a, and panels b and d). Once ATP binds, the filament becomes active (the right side of a and panels c and e).
Figure 3. The structure of the pre‐ and postsynaptic RecA filament. (a) A single turn of the presynaptic RecA filament. RecA monomers are coloured green, cyan and brown; ssDNA is in red and the location of ATP is indicated by the fuchsia Connolly surfaces. (b) The postsynaptic filament with the product complex located approximately in the centre. The incoming strand of DNA is coloured blue. Black lines, filament axis. (c and d) The DNA within the pre‐ and postsynaptic complexes is shown with dsDNA for comparison. In the schematic, the phosphodiester backbones are coloured light green and pale orange, so that the perpendicular arrangement of the bases (coloured purple and blue) is evident. Coordinates for this image were taken from Chen et al. .
Figure 4. The structure of the nucleoprotein filament is conserved. Electron micrographs of nucleoprotein filaments formed on dsDNA. (a) RecA nucleoprotein filament assembled on nicked, circular dsDNA. (b) UvsX nucleoprotein filament assembled on linear M13 dsDNA. (c) Rad51 nucleoprotein filament assembled on nicked, circular ϕX dsDNA. The scale bar in panels (a) and (b) are 0.1 µm. In panel (c), the tobacco mosaic virus particles (indicated by the arrows) are approximately 200 nm in diameter. A more comprehensive version of this figure and legend appears in Bianco et al. .
Figure 5. The DNA‐strand exchange reaction promoted by the RecA protein. RecA is represented as blue spheres and ssDNA‐binding (SSB) protein as orange squares. ssDNA is black and dsDNA is coloured green. The three stages shown are presynapsis (a), synapsis (b) and DNA heteroduplex extension or branch migration (c). Adapted from Tan et al. 2017 © John Wiley and Sons Ltd.
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Further Reading

Bell JC and Kowalczykowski SC (2016a) Mechanics and single‐molecule interrogation of DNA recombination. Annual Review of Biochemistry 85: 193–226.

Bell JC and Kowalczykowski SC (2016b) RecA: regulation and mechanism of a molecular search engine (Trends in Biochemical Sciences, June 2016, Vol. 41, No. 6, 491–507). Trends in Biochemical Sciences 41 (7): 646.

Bianco PR (2010) DNA helicases. EcoSal Plus 4 (1): 1–23.

Cox MM (2007) Regulation of bacterial RecA protein function. Critical Reviews in Biochemistry and Molecular Biology 42 (1): 41–63.

Goodman MF and Woodgate R (2013) Translesion DNA polymerases. Cold Spring Harbor Perspectives in Biology 5 (10): a010363.

Goodman MF, McDonald JP, Jaszczur MM and Woodgate R (2016) Insights into the complex levels of regulation imposed on Escherichia coli DNA polymerase V. DNA Repair (Amst) 44: 42–50.

Kelley WL (2006) Lex marks the spot: the virulent side of SOS and a closer look at the LexA regulon. Molecular Microbiology 62 (5): 1228–1238.

Michel B and Leach D (2012) Homologous recombination‐enzymes and pathways. EcoSal Plus 5 (1): 1–46.

Simmons LA, Foti JJ, Cohen SE and Walker GC (2008) The SOS regulatory network. EcoSal Plus 3 (1): 1–30.

Stasiak A and Di Capua E (1982) The helicity of DNA in complexes with RecA protein. Nature 299: 185–186.

Vaisman A, McDonald JP and Woodgate R (2012) Translesion DNA synthesis. EcoSal Plus 5 (1): 1–32.

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Bianco, Piero R(Mar 2018) RecA Protein. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1002/9780470015902.a0000584.pub3]