Escherichia coli K-12 substr. MG1655 Polypeptide: transcription-repair coupling factor

Gene: mfd Accession Numbers: EG11619 (EcoCyc), b1114, ECK1100

Synonyms: TRCF

Regulation Summary Diagram: ?

Regulation summary diagram for mfd

The Mfd (mutation frequency decline) protein, also known as transcription-repair coupling factor [Selby91], is responsible for ATP-dependent removal of stalled RNA polymerase (RNAP) from DNA lesions [Selby93] by inducing forward movement of the RNA polymerase [Park02] and subsequent recruitment of nucleotide excision repair machinery to the sites of the lesions [Selby93]. In this way Mfd is responsible for preferentially repairing template strand lesions [Selby91]. Mfd discriminates the stalled RNAP-DNA elongation complex and after ATP hydrolysis it disrupts the complex releasing RNAP from the site. Further ATP hydrolysis is required for dissociation of Mfd [Howan12]. Mfd remains bound to DNA in a relatively long-lived complex that may act as a marker for sites of damage [Howan12]. Mfd also rescues roadblocked and back-tracked RNA polymerases by catalyzing the restoration of the forward position [Park02].

The Mfd-dependent transcription coupled repair pathway is able to repair template strand lesions downstream of a stalled RNAP complex. This repair is dependent on the presence of the Mfd protein. Once activated by binding to stalled RNAP, Mfd is able to translocate more than 100 bp along DNA suggesting that repair of downstream lesions is facilitated by the translocation activity of Mfd [Haines14].

Mfd contains helicase motifs with a region similar to RecG involved in binding DNA and ATP hydrolysis, a leucine zipper motif believed to allow binding to RNA polymerase, and a region similar to UvrB which contains a binding domain for UvrA2 [Selby93, Selby95]. The crystal structure of Mfd has been determined to a resolution of 3.2 Å; eight structural domains (D1a - D7) connected by flexible linkers have been identified [Deaconescu06, Deaconescu05]. The crystal structure of the N-terminal region of Mfd has been solved to a resolution of 2.1 Å [Assenmacher06]. The interaction between D2 and D7 maintains Mfd in a 'repressed' conformation in which ATP hydrolysis and DNA translocation are inhibited; derepression is associated with substantial conformational change [Srivastava11]. UvrA binds at the interface of D2 and D7; in the apo enzyme the UvrA binding surface is occluded by the D7 domain; nucleotide binding induces structural rearrangement [Deaconescu12]. In vitro repair assays [Selby91] and UV-induced mutations in vivo [Oller92] show that mfd mutants do not exhibit template strand-specific repair.

Reviews: [Selby94, Roberts04, Deaconescu07, Savery07, Howan14]
Comments: [Van14]

Citations: [Schalow12, Monnet, Ganesan12, Ganesan10, Washburn03, Manelyte10, Murphy09]

Locations: cytosol

Map Position: [1,169,741 <- 1,173,187] (25.21 centisomes, 91°)
Length: 3447 bp / 1148 aa

Molecular Weight of Polypeptide: 129.98 kD (from nucleotide sequence)

Unification Links: ASAP:ABE-0003763 , CGSC:35179 , DIP:DIP-10199N , EchoBASE:EB1576 , EcoGene:EG11619 , EcoliWiki:b1114 , Mint:MINT-1231460 , ModBase:P30958 , OU-Microarray:b1114 , PortEco:mfd , PR:PRO_000023221 , Pride:P30958 , Protein Model Portal:P30958 , RefSeq:NP_415632 , RegulonDB:EG11619 , SMR:P30958 , String:511145.b1114 , Swiss-Model:P30958 , UniProt:P30958

Relationship Links: InterPro:IN-FAMILY:IPR001650 , InterPro:IN-FAMILY:IPR003711 , InterPro:IN-FAMILY:IPR004576 , InterPro:IN-FAMILY:IPR005118 , InterPro:IN-FAMILY:IPR011545 , InterPro:IN-FAMILY:IPR014001 , InterPro:IN-FAMILY:IPR027417 , PDB:Structure:2B2N , PDB:Structure:2EYQ , PDB:Structure:3HJH , PDB:Structure:4DFC , Pfam:IN-FAMILY:PF00270 , Pfam:IN-FAMILY:PF00271 , Pfam:IN-FAMILY:PF02559 , Pfam:IN-FAMILY:PF03461 , Prosite:IN-FAMILY:PS51192 , Prosite:IN-FAMILY:PS51194 , Smart:IN-FAMILY:SM00487 , Smart:IN-FAMILY:SM00490 , Smart:IN-FAMILY:SM00982 , Smart:IN-FAMILY:SM01058

In Paralogous Gene Group: 563 (2 members)

GO Terms:

Biological Process: GO:0000716 - transcription-coupled nucleotide-excision repair, DNA damage recognition Inferred from experiment Inferred by computational analysis [GOA06, Howan12, Selby93]
GO:0006281 - DNA repair Inferred from experiment Inferred by computational analysis [UniProtGOA11a, GOA01a, Selby91]
GO:0006355 - regulation of transcription, DNA-templated Inferred from experiment Inferred by computational analysis [GOA06, Selby93]
GO:0006974 - cellular response to DNA damage stimulus Inferred from experiment Inferred by computational analysis [UniProtGOA11a, Selby93]
Molecular Function: GO:0003677 - DNA binding Inferred from experiment Inferred by computational analysis [UniProtGOA11a, GOA06, Selby95]
GO:0005515 - protein binding Inferred from experiment [Deaconescu12]
GO:0008026 - ATP-dependent helicase activity Inferred from experiment [Selby93]
GO:0015616 - DNA translocase activity Inferred from experiment [Haines14]
GO:0000166 - nucleotide binding Inferred by computational analysis [UniProtGOA11a]
GO:0003676 - nucleic acid binding Inferred by computational analysis [GOA01a]
GO:0003684 - damaged DNA binding Inferred by computational analysis [GOA01a]
GO:0005524 - ATP binding Inferred by computational analysis [UniProtGOA11a, GOA06, GOA01a]
GO:0016787 - hydrolase activity Inferred by computational analysis [UniProtGOA11a, GOA06]
Cellular Component: GO:0005829 - cytosol Inferred from experiment Inferred by computational analysis [DiazMejia09, Ishihama08]
GO:0005737 - cytoplasm Inferred by computational analysis [UniProtGOA11, UniProtGOA11a, GOA06]

MultiFun Terms: information transfer RNA related Transcription related

Essentiality data for mfd 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]

Last-Curated ? 20-Apr-2015 by Mackie A , Macquarie University

Sequence Features

Protein sequence of transcription-repair coupling factor with features indicated

Feature Class Location Common Name Homology Motif Citations Comment
Protein-Structure-Region 50 -> 117 D1a  
D1a/D2/D1b form the UvrB homology module
Protein-Structure-Region 118 -> 216 D2  
D1a/D2/D1b form the UvrB homology module
Protein-Structure-Region 217 -> 340 D1b  
D1a/D2/D1b form the UvrB homology module
Protein-Structure-Region 353 -> 454 D3  
non conserved species specific domain
Sequence-Conflict 365    
[Selby93, UniProt10a]
UniProt: (in Ref. 1);
Protein-Structure-Region 478 -> 545 D4  
[Deaconescu06, Deaconescu06]
RNAP interacting domain (RID); L499R substitution results in mutant protein that is able to bind DNA and hydrolyse ATP but is unable to displace RNA polymerase
Protein-Structure-Region 547 -> 780 D5  
translocation domain 1
Conserved-Region 615 -> 776    
UniProt: Helicase ATP-binding;
Nucleotide-Phosphate-Binding-Region 628 -> 635 Walker A GXXXXGK(T/S)
[UniProt10, Deaconescu06]
UniProt: ATP; Non-Experimental Qualifier: potential;
Conserved-Region 724 -> 732 Walker B  
Protein-Segment 729 -> 732    
UniProt: DEEH box; Sequence Annotation Type: short sequence motif.
Protein-Structure-Region 781 -> 990 D6  
translocation domain 2
Conserved-Region 798 -> 951    
UniProt: Helicase C-terminal;
Conserved-Region 926 -> 965 TRG motif  
conserved between RecG and TRCF families (translocation in RecG motif); essential for RNAP release
Mutagenesis-Variant 929    
R → A: impairs RNAP displacement activity in vitro, no effect on DNA binding / ATP hydrolysis
Mutagenesis-Variant 948    
H → A: impairs RNAP displacement activity in vitro, no effect on DNA binding / ATP hydrolysis
Mutagenesis-Variant 953    
R → A: impairs RNAP displacement activity in vitro, no effect on DNA binding / ATP hydrolysis
Mutagenesis-Variant 963    
Q → A: impairs RNAP displacement activity in vitro, no effect on DNA binding / ATP hydrolysis
Protein-Structure-Region 1002 -> 1148 D7  

Gene Local Context (not to scale): ?

Gene local context diagram

Transcription Units:

Transcription-unit diagram

Transcription-unit diagram


10/20/97 Gene b1114 from Blattner lab Genbank (v. M52) entry merged into EcoCyc gene EG11619; confirmed by SwissProt match.


Assenmacher06: Assenmacher N, Wenig K, Lammens A, Hopfner KP (2006). "Structural Basis for Transcription-coupled Repair: the N Terminus of Mfd Resembles UvrB with Degenerate ATPase Motifs." J Mol Biol 355(4):675-83. PMID: 16309703

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

Chambers03: Chambers AL, Smith AJ, Savery NJ (2003). "A DNA translocation motif in the bacterial transcription--repair coupling factor, Mfd." Nucleic Acids Res 31(22);6409-18. PMID: 14602898

Deaconescu05: Deaconescu AM, Darst SA (2005). "Crystallization and preliminary structure determination of Escherichia coli Mfd, the transcription-repair coupling factor." Acta Crystallograph Sect F Struct Biol Cryst Commun 61(Pt 12);1062-4. PMID: 16511235

Deaconescu06: Deaconescu AM, Chambers AL, Smith AJ, Nickels BE, Hochschild A, Savery NJ, Darst SA (2006). "Structural basis for bacterial transcription-coupled DNA repair." Cell 124(3);507-20. PMID: 16469698

Deaconescu07: Deaconescu AM, Savery N, Darst SA (2007). "The bacterial transcription repair coupling factor." Curr Opin Struct Biol 17(1);96-102. PMID: 17239578

Deaconescu12: Deaconescu AM, Sevostyanova A, Artsimovitch I, Grigorieff N (2012). "Nucleotide excision repair (NER) machinery recruitment by the transcription-repair coupling factor involves unmasking of a conserved intramolecular interface." Proc Natl Acad Sci U S A 109(9);3353-8. PMID: 22331906

DiazMejia09: Diaz-Mejia JJ, Babu M, Emili A (2009). "Computational and experimental approaches to chart the Escherichia coli cell-envelope-associated proteome and interactome." FEMS Microbiol Rev 33(1);66-97. PMID: 19054114

Ganesan10: Ganesan AK, Hanawalt PC (2010). "Transcription-coupled nucleotide excision repair of a gene transcribed by bacteriophage T7 RNA polymerase in Escherichia coli." DNA Repair (Amst) 9(9);958-63. PMID: 20638914

Ganesan12: Ganesan A, Spivak G, Hanawalt PC (2012). "Transcription-coupled DNA repair in prokaryotes." Prog Mol Biol Transl Sci 110;25-40. PMID: 22749141

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

GOA01a: 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."

Haines14: Haines NM, Kim YI, Smith AJ, Savery NJ (2014). "Stalled transcription complexes promote DNA repair at a distance." Proc Natl Acad Sci U S A 111(11);4037-42. PMID: 24554077

Howan12: Howan K, Smith AJ, Westblade LF, Joly N, Grange W, Zorman S, Darst SA, Savery NJ, Strick TR (2012). "Initiation of transcription-coupled repair characterized at single-molecule resolution." Nature 490(7420);431-4. PMID: 22960746

Howan14: Howan K, Monnet J, Fan J, Strick TR (2014). "Stopped in its tracks: the RNA polymerase molecular motor as a robust sensor of DNA damage." DNA Repair (Amst) 20;49-57. PMID: 24685770

Ishihama08: Ishihama Y, Schmidt T, Rappsilber J, Mann M, Hartl FU, Kerner MJ, Frishman D (2008). "Protein abundance profiling of the Escherichia coli cytosol." BMC Genomics 9;102. PMID: 18304323

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

Manelyte10: Manelyte L, Kim YI, Smith AJ, Smith RM, Savery NJ (2010). "Regulation and rate enhancement during transcription-coupled DNA repair." Mol Cell 40(5);714-24. PMID: 21145481

Monnet: Monnet J, Grange W, Strick TR, Joly N "Mfd as a central partner of transcription coupled repair." Transcription 4(3);109-13. PMID: 23756341

Murphy09: Murphy MN, Gong P, Ralto K, Manelyte L, Savery NJ, Theis K (2009). "An N-terminal clamp restrains the motor domains of the bacterial transcription-repair coupling factor Mfd." Nucleic Acids Res 37(18);6042-53. PMID: 19700770

Oller92: Oller AR, Fijalkowska IJ, Dunn RL, Schaaper RM (1992). "Transcription-repair coupling determines the strandedness of ultraviolet mutagenesis in Escherichia coli." Proc Natl Acad Sci U S A 89(22);11036-40. PMID: 1438310

Park02: Park JS, Marr MT, Roberts JW (2002). "E. coli Transcription repair coupling factor (Mfd protein) rescues arrested complexes by promoting forward translocation." Cell 109(6);757-67. PMID: 12086674

Roberts04: Roberts J, Park JS (2004). "Mfd, the bacterial transcription repair coupling factor: translocation, repair and termination." Curr Opin Microbiol 7(2);120-5. PMID: 15063847

Savery07: Savery NJ (2007). "The molecular mechanism of transcription-coupled DNA repair." Trends Microbiol 15(7);326-33. PMID: 17572090

Schalow12: Schalow BJ, Courcelle CT, Courcelle J (2012). "Mfd is required for rapid recovery of transcription following UV-induced DNA damage but not oxidative DNA damage in Escherichia coli." J Bacteriol 194(10);2637-45. PMID: 22427630

Selby91: Selby CP, Witkin EM, Sancar A (1991). "Escherichia coli mfd mutant deficient in "mutation frequency decline" lacks strand-specific repair: in vitro complementation with purified coupling factor." Proc Natl Acad Sci U S A 88(24);11574-8. PMID: 1763073

Selby93: Selby CP, Sancar A (1993). "Molecular mechanism of transcription-repair coupling." Science 260(5104);53-8. PMID: 8465200

Selby94: Selby CP, Sancar A (1994). "Mechanisms of transcription-repair coupling and mutation frequency decline." Microbiol Rev 58(3);317-29. PMID: 7968917

Selby95: Selby CP, Sancar A (1995). "Structure and function of transcription-repair coupling factor. I. Structural domains and binding properties." J Biol Chem 270(9);4882-9. PMID: 7876261

Srivastava11: Srivastava DB, Darst SA (2011). "Derepression of bacterial transcription-repair coupling factor is associated with a profound conformational change." J Mol Biol 406(2);275-84. PMID: 21185303

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

UniProt10: UniProt Consortium (2010). "UniProt version 2010-07 released on 2010-06-15 00:00:00." Database.

UniProt10a: UniProt Consortium (2010). "UniProt version 2010-11 released on 2010-11-02 00:00:00." Database.

UniProt15: UniProt Consortium (2015). "UniProt version 2015-01 released on 2015-01-16 00:00:00." Database.

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

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

Van14: Van Houten B, Kisker C (2014). "Transcriptional pausing to scout ahead for DNA damage." Proc Natl Acad Sci U S A 111(11);3905-6. PMID: 24599593

Washburn03: Washburn RS, Wang Y, Gottesman ME (2003). "Role of E.coli transcription-repair coupling factor Mfd in Nun-mediated transcription termination." J Mol Biol 329(4);655-62. PMID: 12787667

Other References Related to Gene Regulation

Ogasawara05: Ogasawara H, Teramoto J, Yamamoto S, Hirao K, Yamamoto K, Ishihama A, Utsumi R (2005). "Negative regulation of DNA repair gene (uvrA) expression by ArcA/ArcB two-component system in Escherichia coli." FEMS Microbiol Lett 251(2);243-9. PMID: 16140472

Stanley03: Stanley LK, Savery NJ (2003). "Characterisation of the Escherichia coli mfd promoter." Arch Microbiol 2003;179(5);381-5. PMID: 12658334

Wade06: Wade JT, Roa DC, Grainger DC, Hurd D, Busby SJ, Struhl K, Nudler E (2006). "Extensive functional overlap between sigma factors in Escherichia coli." Nat Struct Mol Biol 13(9);806-14. PMID: 16892065

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Please cite the following article in publications resulting from the use of EcoCyc: Nucleic Acids Research 41:D605-12 2013
Page generated by SRI International Pathway Tools version 19.0 on Fri Oct 9, 2015, biocyc13.