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Escherichia coli K-12 substr. MG1655 Polypeptide: DNA strand exchange and recombination protein with protease and nuclease activity



Gene: recA Accession Numbers: EG10823 (EcoCyc), b2699, ECK2694

Synonyms: zab, umuB, tif, lexB, recH, rnmB, srf

Regulation Summary Diagram: ?

Summary:
In Escherichia coli K-12, RecA serves a variety of biochemical functions: RecA functions in homologous recombination by catalyzing DNA strand exchange reactions; it serves as a regulatory protein to induce the SOS response to DNA damage by its action on the repressor protein LexA; and it is a required component for the mutagenic bypass of DNA lesions during the SOS response [Kuzminov99, Courcelle03].

recA is not a part of any operon and it is surrounded by genes unrelated to DNA metabolism. Normally, recA expression is such that constitutive levels of RecA are maintained at between 1,000 and 10,000 monomers per cell but is induced by DNA damage, e.g. UV irradiation, to 50 times its normal level [Karu82].

DNA damage can occur due to a variety of environmental assaults including UV irradiation and chemical agents. E. coli has a number of complex enzymatic pathways for the repair of DNA damage (see DNA Repair Enzymes).

In most cases, DNA damage is on only 1 of 2 strands, but in some cases damage can occur to both strands. Double strand breaks (DSBs) cannot be repaired via excision repair pathways because by their nature they lack an intact strand to be used as a template. Repair of DSBs utilizes the RecA pathway of homologous recombination with a separate, intact, homologous region of DNA.

One of the roles of RecA is in repair of DSBs by homologous recombination in which a separate intact region of DNA is used. Studies in which double-strand breaks are induced in vivo in E.coli DNA show that double-strand break repair (DSBR) can proceed via one of two recombination pathways. Both pathways require RecBCD and RecA, but one depends on the resolvase enzyme, RuvABC, while the other does not and instead relies on RecG [Meddows04]. RecA's role in both pathways is to catalyze homologous recombination. RecA polymerizes to form filaments around both single-strand and double-strand DNA and in doing so extends the DNA within the filament to 1.5 times its unbound length [Courcelle03]. Filamented RecA depolymerizes readily, and is able to generate torsion in DNA even when the ends of the DNA are able to spin freely [Fulconis06]. RecA filament nucleation requires ATP binding, but not hydrolysis, whereas subsequent bidirectional filament growth depends on neither [Galletto06]. The RecA filament has two DNA-binding sites, one of which binds to single stranded DNA and the other binds duplex DNA. RecA performs a homology search of the duplex DNA until a homologous strand is located. The single-strand DNA is paired with homologous duplex DNA in a reaction that yields a RecA protein filament bound in a three or four-stranded DNA structure (Holliday junction) [Courcelle03]. The recombination junction is then resolved through either the actions of the RuvABC resolvasome or RecG helicase. Both pathways catalyze branch migration on recombinational junctions and the resolution of the junction after recombination [Courcelle03, Whitby94, Parsons92].

In RecA's role in the SOS response, RecA binds to single stranded regions generated at regions of stalled replication and the bound RecA becomes activated. The activated RecA promotes the autocatalytic cleavage of three proteins: the SOS response repressor (LexA), the lambda phage repressor and the UmuD protein. Cleavage of the lambda phage repressor plays no role in DNA repair, but stimulates prophage induction. Cleavage of the UmuD protein creates the cleavage product, UmuD', which then combines with UmuC to form a complex, UmuD'C, also known as polymerase V, which plays a role in catalyzing translesion DNA synthesis, an alternative to repair which inserts random bases opposite some noncoding lesions. This strategy can result in point mutations, but the error-prone synthesis may be a life-saving option in cases of massive DNA damage [Kuzminov99, Rajagopalan92]. Activated RecA becomes a regulator of the SOS response by inducing the autocatalytic cleavage of LexA. LexA binds to a 20 base pair consensus sequence in the operator region of most of the DNA damage-inducible genes and suppresses their expression. As the cellular concentration of LexA decreases, the SOS genes become more frequently transcribed. These genes include uvrA, B and C, the nucleotide excision repair genes polB, dinB and umuC and D, which are genes coding for DNA polymerases II, IV and V, respectively. lexA and recA are also derepressed in the SOS response. Also, sfiA, which inhibits cell division, is derepressed - presumably allowing time for DNA repair to take place.

RecA filament assembly has been visualized in vitro on single molecules of SSB (single strand binding protein)-coated ssDNA - the relevant in vivo substrate. Filament nucleation requires a RecG dimer, filament growth occurs by monomer addition and is bidirectional in nature. RecA filaments grow at approximately 30-300 monomers/minute. RecA nucleation is stimulated by the introduction of RecF whilst filament nucleation and growth are stimulated by the addition of the mediator proteins RecO and RecR [Bell12]. Nucleation of RecA onto ssDNA ends triggers polymerisation of RecA into bundles which are critical for homologous recombination repair between distant regions of homolgy. RecA bundles are estimated to be 1-4 µM in length and contain approximately 70% of the normal cellular pool of RecA molecules. The majority of RecA molecules in these bundles do not directly contact DNA [Lesterlin14].

The binding of RecA to single-stranded DNA in regions of stalled replication also serves a structural role - protecting ssDNA from degradation by exonucleases while awaiting repair [Courcelle03a]. Mutation deletion studies [Courcelle97] show that the resumption of replication at DNA replication forks also requires the presence of the recF and recR gene products.

The X-ray crystal structure of uncomplexed E. coli RecA protein has been determined in three new crystal forms at resolutions of 1.9 Å, 2.0 Å, and 2.6 Å [Xing04].

recA is one of a network of 93 genes believed to play a role in promoting the stress-induced mutagenesis (SIM) response of E. coli K-12 [Al12].

Non-synonymous point mutations in recA were identified in cell populations that were selected for high resistance to ionizing radiation. The D276A allele was tested and was found to contribute significantly to resistance [Byrne14a].

Citations: [Lee09, Lovett12]

Gene Citations: [Pages03, Stohl03, Clark94, Fernandez00]

Locations: cytosol

Map Position: [2,820,730 <- 2,821,791] (60.8 centisomes)
Length: 1062 bp / 353 aa

Molecular Weight of Polypeptide: 37.973 kD (from nucleotide sequence)

Unification Links: ASAP:ABE-0008876 , CGSC:312 , DIP:DIP-31832N , EchoBASE:EB0816 , EcoGene:EG10823 , EcoliWiki:b2699 , Mint:MINT-1300726 , OU-Microarray:b2699 , PortEco:recA , PR:PRO_000023696 , Pride:P0A7G6 , Protein Model Portal:P0A7G6 , RefSeq:NP_417179 , RegulonDB:EG10823 , SMR:P0A7G6 , String:511145.b2699 , UniProt:P0A7G6

Relationship Links: InterPro:IN-FAMILY:IPR003593 , InterPro:IN-FAMILY:IPR013765 , InterPro:IN-FAMILY:IPR020584 , InterPro:IN-FAMILY:IPR020587 , InterPro:IN-FAMILY:IPR020588 , InterPro:IN-FAMILY:IPR023400 , InterPro:IN-FAMILY:IPR027417 , Panther:IN-FAMILY:PTHR22942:SF1 , PDB:Structure:1AA3 , PDB:Structure:1N03 , PDB:Structure:1REA , PDB:Structure:1U94 , PDB:Structure:1U98 , PDB:Structure:1U99 , PDB:Structure:1XMS , PDB:Structure:1XMV , PDB:Structure:2REB , PDB:Structure:2REC , PDB:Structure:3CMT , PDB:Structure:3CMU , PDB:Structure:3CMV , PDB:Structure:3CMW , PDB:Structure:3CMX , Pfam:IN-FAMILY:PF00154 , Prints:IN-FAMILY:PR00142 , Prosite:IN-FAMILY:PS00321 , Prosite:IN-FAMILY:PS50162 , Prosite:IN-FAMILY:PS50163 , Smart:IN-FAMILY:SM00382

In Paralogous Gene Group: 445 (2 members)

Genetic Regulation Schematic: ?

GO Terms:

Biological Process: GO:0006310 - DNA recombination Inferred from experiment Inferred by computational analysis [UniProtGOA11, GOA06, Adikesavan11]
GO:0006974 - cellular response to DNA damage stimulus Inferred from experiment Inferred by computational analysis [UniProtGOA11, Khil02, Fernandez00]
GO:0009432 - SOS response Inferred from experiment Inferred by computational analysis [UniProtGOA11, GOA06, GOA01, Vlaši11, Witkin74]
GO:0010212 - response to ionizing radiation Inferred from experiment [Byrne14a]
GO:0048870 - cell motility Inferred from experiment [GomezGomez07]
GO:0006200 - ATP catabolic process Inferred by computational analysis [GOA06, GOA01]
GO:0006259 - DNA metabolic process Inferred by computational analysis [GOA01]
GO:0006281 - DNA repair Inferred by computational analysis [UniProtGOA11, GOA06, GOA01]
GO:0006950 - response to stress Inferred by computational analysis [UniProtGOA11]
Molecular Function: GO:0003697 - single-stranded DNA binding Inferred from experiment Inferred by computational analysis [GOA06, GOA01, Roy09]
GO:0005515 - protein binding Inferred from experiment [Zhang12a]
GO:0000166 - nucleotide binding Inferred by computational analysis [UniProtGOA11]
GO:0003677 - DNA binding Inferred by computational analysis [UniProtGOA11, GOA01]
GO:0003684 - damaged DNA binding Inferred by computational analysis [GOA06]
GO:0005524 - ATP binding Inferred by computational analysis [UniProtGOA11, GOA06, GOA01]
GO:0008094 - DNA-dependent ATPase activity Inferred by computational analysis [GOA06, GOA01]
Cellular Component: GO:0005737 - cytoplasm Inferred by computational analysis [UniProtGOA11a, UniProtGOA11, GOA06]
GO:0005829 - cytosol

MultiFun Terms: cell processes SOS response
information transfer DNA related DNA recombination
information transfer DNA related DNA repair
information transfer protein related turnover, degradation
regulation type of regulation posttranscriptional proteases, cleavage of compounds

Essentiality data for recA knockouts: ?

Growth Medium Growth? T (°C) O2 pH Osm/L Growth Observations
LB enriched Yes 37 Aerobic 6.95   Yes [Gerdes03, Comment 1]
LB Lennox Yes 37 Aerobic 7   Yes [Baba06, Comment 2]
M9 medium with 1% glycerol Yes 37 Aerobic 7.2 0.35 Yes [Joyce06, Comment 3]
MOPS medium with 0.4% glucose Yes 37 Aerobic 7.2 0.22 Yes [Baba06, Comment 2]

Sequence Features

Feature Class Location Citations Comment
Cleavage-of-Initial-Methionine 1
[Horii80, Sancar80, UniProt11]
UniProt: Removed.
Chain 2 -> 353
[UniProt09]
UniProt: Protein recA;
Nucleotide-Phosphate-Binding-Region 67 -> 74
[UniProt10]
UniProt: ATP;
Sequence-Conflict 113
[Yamamoto97a, UniProt10]
Alternate sequence: D → E; UniProt: (in Ref. 4);
Sequence-Conflict 191
[Gardner95, UniProt10]
Alternate sequence: I → missing; UniProt: (in Ref. 8; AA sequence);
Acetylation-Modification 287
[Yu08]
 


Gene Local Context (not to scale): ?

Transcription Units:

Notes:

History:
10/20/97 Gene b2699 from Blattner lab Genbank (v. M52) entry merged into EcoCyc gene EG10823; confirmed by SwissProt match.


References

Adikesavan11: Adikesavan AK, Katsonis P, Marciano DC, Lua R, Herman C, Lichtarge O (2011). "Separation of recombination and SOS response in Escherichia coli RecA suggests LexA interaction sites." PLoS Genet 7(9);e1002244. PMID: 21912525

Al12: Al Mamun AA, Lombardo MJ, Shee C, Lisewski AM, Gonzalez C, Lin D, Nehring RB, Saint-Ruf C, Gibson JL, Frisch RL, Lichtarge O, Hastings PJ, Rosenberg SM (2012). "Identity and function of a large gene network underlying mutagenic repair of DNA breaks." Science 338(6112);1344-8. PMID: 23224554

Baba06: Baba T, Ara T, Hasegawa M, Takai Y, Okumura Y, Baba M, Datsenko KA, Tomita M, Wanner BL, Mori H (2006). "Construction of Escherichia coli K-12 in-frame, single-gene knockout mutants: the Keio collection." Mol Syst Biol 2;2006.0008. PMID: 16738554

Bell12: Bell JC, Plank JL, Dombrowski CC, Kowalczykowski SC (2012). "Direct imaging of RecA nucleation and growth on single molecules of SSB-coated ssDNA." Nature 491(7423);274-8. PMID: 23103864

Byrne14a: Byrne RT, Klingele AJ, Cabot EL, Schackwitz WS, Martin JA, Martin J, Wang Z, Wood EA, Pennacchio C, Pennacchio LA, Perna NT, Battista JR, Cox MM (2014). "Evolution of extreme resistance to ionizing radiation via genetic adaptation of DNA repair." Elife 3;e01322. PMID: 24596148

Clark94: Clark AJ, Sandler SJ (1994). "Homologous genetic recombination: the pieces begin to fall into place." Crit Rev Microbiol 1994;20(2);125-42. PMID: 8080625

Courcelle03: Courcelle J, Hanawalt PC (2003). "RecA-dependent recovery of arrested DNA replication forks." Annu Rev Genet 37;611-46. PMID: 14616075

Courcelle03a: Courcelle J, Donaldson JR, Chow KH, Courcelle CT (2003). "DNA damage-induced replication fork regression and processing in Escherichia coli." Science 299(5609);1064-7. PMID: 12543983

Courcelle97: Courcelle J, Carswell-Crumpton C, Hanawalt PC (1997). "recF and recR are required for the resumption of replication at DNA replication forks in Escherichia coli." Proc Natl Acad Sci U S A 94(8);3714-9. PMID: 9108043

Emmerson68: Emmerson PT (1968). "Recombination deficient mutants of Escherichia coli K12 that map between thy A and argA." Genetics 60(1);19-30. PMID: 4884588

Fernandez00: Fernandez De Henestrosa AR, Ogi T, Aoyagi S, Chafin D, Hayes JJ, Ohmori H, Woodgate R (2000). "Identification of additional genes belonging to the LexA regulon in Escherichia coli." Mol Microbiol 35(6);1560-72. PMID: 10760155

Fulconis06: Fulconis R, Mine J, Bancaud A, Dutreix M, Viovy JL (2006). "Mechanism of RecA-mediated homologous recombination revisited by single molecule nanomanipulation." EMBO J 25(18);4293-304. PMID: 16946710

Galletto06: Galletto R, Amitani I, Baskin RJ, Kowalczykowski SC (2006). "Direct observation of individual RecA filaments assembling on single DNA molecules." Nature 443(7113);875-8. PMID: 16988658

Gardner95: Gardner RV, Voloshin ON, Camerini-Otero RD (1995). "The identification of the single-stranded DNA-binding domain of the Escherichia coli RecA protein." Eur J Biochem 233(2);419-25. PMID: 7588783

Gerdes03: Gerdes SY, Scholle MD, Campbell JW, Balazsi G, Ravasz E, Daugherty MD, Somera AL, Kyrpides NC, Anderson I, Gelfand MS, Bhattacharya A, Kapatral V, D'Souza M, Baev MV, Grechkin Y, Mseeh F, Fonstein MY, Overbeek R, Barabasi AL, Oltvai ZN, Osterman AL (2003). "Experimental determination and system level analysis of essential genes in Escherichia coli MG1655." J Bacteriol 185(19);5673-84. PMID: 13129938

GOA01: GOA, DDB, FB, MGI, ZFIN (2001). "Gene Ontology annotation through association of InterPro records with GO terms."

GOA06: GOA, SIB (2006). "Electronic Gene Ontology annotations created by transferring manual GO annotations between orthologous microbial proteins."

GomezGomez07: Gomez-Gomez JM, Manfredi C, Alonso JC, Blazquez J (2007). "A novel role for RecA under non-stress: promotion of swarming motility in Escherichia coli K-12." BMC Biol 5;14. PMID: 17391508

Horii80: Horii T, Ogawa T, Ogawa H (1980). "Organization of the recA gene of Escherichia coli." Proc Natl Acad Sci U S A 77(1);313-7. PMID: 6244554

Joyce06: Joyce AR, Reed JL, White A, Edwards R, Osterman A, Baba T, Mori H, Lesely SA, Palsson BO, Agarwalla S (2006). "Experimental and computational assessment of conditionally essential genes in Escherichia coli." J Bacteriol 188(23);8259-71. PMID: 17012394

Karu82: Karu AE, Belk ED (1982). "Induction of E. coli recA protein via recBC and alternate pathways: quantitation by enzyme-linked immunosorbent assay (ELISA)." Mol Gen Genet 185(2);275-82. PMID: 6283318

Khil02: Khil PP, Camerini-Otero RD (2002). "Over 1000 genes are involved in the DNA damage response of Escherichia coli." Mol Microbiol 44(1);89-105. PMID: 11967071

Kuzminov99: Kuzminov A (1999). "Recombinational repair of DNA damage in Escherichia coli and bacteriophage lambda." Microbiol Mol Biol Rev 63(4);751-813, table of contents. PMID: 10585965

Lee09: Lee CD, Wang TF (2009). "The N-terminal domain of Escherichia coli RecA have multiple functions in promoting homologous recombination." J Biomed Sci 16;37. PMID: 19338667

Lesterlin14: Lesterlin C, Ball G, Schermelleh L, Sherratt DJ (2014). "RecA bundles mediate homology pairing between distant sisters during DNA break repair." Nature 506(7487);249-53. PMID: 24362571

Lovett12: Lovett ST (2012). "Biochemistry: A glimpse of molecular competition." Nature 491(7423);198-200. PMID: 23103870

Meddows04: Meddows TR, Savory AP, Lloyd RG (2004). "RecG helicase promotes DNA double-strand break repair." Mol Microbiol 52(1);119-32. PMID: 15049815

Pages03: Pages V, Koffel-Schwartz N, Fuchs RP (2003). "recX, a new SOS gene that is co-transcribed with the recA gene in Escherichia coli." DNA Repair (Amst) 2003;2(3);273-84. PMID: 12547390

Parsons92: Parsons CA, Tsaneva I, Lloyd RG, West SC (1992). "Interaction of Escherichia coli RuvA and RuvB proteins with synthetic Holliday junctions." Proc Natl Acad Sci U S A 89(12);5452-6. PMID: 1608954

Rajagopalan92: Rajagopalan M, Lu C, Woodgate R, O'Donnell M, Goodman MF, Echols H (1992). "Activity of the purified mutagenesis proteins UmuC, UmuD', and RecA in replicative bypass of an abasic DNA lesion by DNA polymerase III." Proc Natl Acad Sci U S A 89(22);10777-81. PMID: 1438275

Roy09: Roy R, Kozlov AG, Lohman TM, Ha T (2009). "SSB protein diffusion on single-stranded DNA stimulates RecA filament formation." Nature 461(7267);1092-7. PMID: 19820696

Sancar80: Sancar A, Stachelek C, Konigsberg W, Rupp WD (1980). "Sequences of the recA gene and protein." Proc Natl Acad Sci U S A 77(5);2611-5. PMID: 6930655

Stohl03: Stohl EA, Brockman JP, Burkle KL, Morimatsu K, Kowalczykowski SC, Seifert HS (2003). "Escherichia coli RecX inhibits RecA recombinase and coprotease activities in vitro and in vivo." J Biol Chem 2003;278(4);2278-85. PMID: 12427742

UniProt09: UniProt Consortium (2009). "UniProt version 15.8 released on 2009-10-01 00:00:00." Database.

UniProt10: UniProt Consortium (2010). "UniProt version 2010-11 released on 2010-11-02 00:00:00." Database.

UniProt11: UniProt Consortium (2011). "UniProt version 2011-06 released on 2011-06-30 00:00:00." Database.

UniProtGOA11: UniProt-GOA (2011). "Gene Ontology annotation based on manual assignment of UniProtKB keywords in UniProtKB/Swiss-Prot entries."

UniProtGOA11a: UniProt-GOA (2011). "Gene Ontology annotation based on the manual assignment of UniProtKB Subcellular Location terms in UniProtKB/Swiss-Prot entries."

Vlaši11: Vlašić I, Šimatović A, Brčić-Kostić K (2011). "Genetic requirements for high constitutive SOS expression in recA730 mutants of Escherichia coli." J Bacteriol 193(18);4643-51. PMID: 21764927

Whitby94: Whitby MC, Vincent SD, Lloyd RG (1994). "Branch migration of Holliday junctions: identification of RecG protein as a junction specific DNA helicase." EMBO J 13(21);5220-8. PMID: 7957087

Witkin74: Witkin EM (1974). "Thermal enhancement of ultraviolet mutability in a tif-1 uvrA derivative of Escherichia coli B-r: evidence that ultraviolet mutagenesis depends upon an inducible function." Proc Natl Acad Sci U S A 71(5);1930-4. PMID: 4600265

Xing04: Xing X, Bell CE (2004). "Crystal structures of Escherichia coli RecA in a compressed helical filament." J Mol Biol 342(5);1471-85. PMID: 15364575

Yamamoto97a: Yamamoto Y, Aiba H, Baba T, Hayashi K, Inada T, Isono K, Itoh T, Kimura S, Kitagawa M, Makino K, Miki T, Mitsuhashi N, Mizobuchi K, Mori H, Nakade S, Nakamura Y, Nashimoto H, Oshima T, Oyama S, Saito N, Sampei G, Satoh Y, Sivasundaram S, Tagami H, Horiuchi T (1997). "Construction of a contiguous 874-kb sequence of the Escherichia coli -K12 genome corresponding to 50.0-68.8 min on the linkage map and analysis of its sequence features." DNA Res 4(2);91-113. PMID: 9205837

Yu08: Yu BJ, Kim JA, Moon JH, Ryu SE, Pan JG (2008). "The diversity of lysine-acetylated proteins in Escherichia coli." J Microbiol Biotechnol 18(9);1529-36. PMID: 18852508

Zhang12a: Zhang M, Li T, Wang H, Cheng F, Zhou Y, Bi L, Zhang XE (2012). "MutL associates with Escherichia coli RecA and inhibits its ATPase activity." Arch Biochem Biophys 517(2);98-103. PMID: 22001225

Other References Related to Gene Regulation

Brent81: Brent R, Ptashne M (1981). "Mechanism of action of the lexA gene product." Proc Natl Acad Sci U S A 1981;78(7);4204-8. PMID: 7027256

Little81: Little JW, Mount DW, Yanisch-Perron CR (1981). "Purified lexA protein is a repressor of the recA and lexA genes." Proc Natl Acad Sci U S A 1981;78(7);4199-203. PMID: 7027255


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Please cite the following article in publications resulting from the use of EcoCyc: Nucleic Acids Research 41:D605-12 2013
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