TRFA_ECOLX
ID TRFA_ECOLX Reviewed; 382 AA.
AC P07676;
DT 01-APR-1988, integrated into UniProtKB/Swiss-Prot.
DT 01-APR-1988, sequence version 1.
DT 25-MAY-2022, entry version 65.
DE RecName: Full=Plasmid replication initiator protein TrfA;
GN Name=trfA;
OS Escherichia coli.
OG Plasmid IncP-alpha RK2.
OC Bacteria; Proteobacteria; Gammaproteobacteria; Enterobacterales;
OC Enterobacteriaceae; Escherichia.
OX NCBI_TaxID=562;
RN [1]
RP NUCLEOTIDE SEQUENCE [GENOMIC DNA].
RC PLASMID=IncP-alpha RK2;
RX PubMed=6374158; DOI=10.1016/0022-2836(84)90347-4;
RA Smith C.A., Thomas C.M.;
RT "Nucleotide sequence of the trfA gene of broad host-range plasmid RK2.";
RL J. Mol. Biol. 175:251-262(1984).
RN [2]
RP SUBUNIT, AND DNA-BINDING.
RC PLASMID=IncP-alpha RK2;
RX PubMed=8636140; DOI=10.1074/jbc.271.12.7072;
RA Toukdarian A.E., Helinski D.R., Perri S.;
RT "The plasmid RK2 initiation protein binds to the origin of replication as a
RT monomer.";
RL J. Biol. Chem. 271:7072-7078(1996).
RN [3]
RP FUNCTION IN PLASMID COPY NUMBER, DNA-BINDING, AND MUTAGENESIS OF GLY-254
RP AND SER-267.
RC PLASMID=IncP-alpha RK2;
RX PubMed=8622975; DOI=10.1073/pnas.93.8.3559;
RA Blasina A., Kittell B.L., Toukdarian A.E., Helinski D.R.;
RT "Copy-up mutants of the plasmid RK2 replication initiation protein are
RT defective in coupling RK2 replication origins.";
RL Proc. Natl. Acad. Sci. U.S.A. 93:3559-3564(1996).
RN [4]
RP FUNCTION IN DNA REPLICATION, DNA-BINDING, AND MUTAGENESIS OF GLY-254 AND
RP SER-267.
RC PLASMID=IncP-alpha RK2;
RX PubMed=9242693; DOI=10.1074/jbc.272.32.20173;
RA Konieczny I., Doran K.S., Helinski D.R., Blasina A.;
RT "Role of TrfA and DnaA proteins in origin opening during initiation of DNA
RT replication of the broad host range plasmid RK2.";
RL J. Biol. Chem. 272:20173-20178(1997).
RN [5]
RP SUBCELLULAR LOCATION.
RC STRAIN=CSR603; PLASMID=IncP-alpha RK2;
RX PubMed=10692384; DOI=10.1128/jb.182.6.1757-1760.2000;
RA Kim P.D., Firshein W.;
RT "Isolation of an inner membrane-derived subfraction that supports in vitro
RT replication of a mini-RK2 plasmid in Escherichia coli.";
RL J. Bacteriol. 182:1757-1760(2000).
RN [6]
RP SUBCELLULAR LOCATION, TOXIC FRAGMENT, AND MUTAGENESIS OF 286-VAL--ALA-297;
RP 289-ASP--GLU-291; 293-VAL--VAL-294 AND PHE-296.
RC STRAIN=K12 / DH5-alpha; PLASMID=IncP-alpha RK2;
RX PubMed=10783300; DOI=10.1006/plas.2000.1467;
RA Kim P.D., Rosche T.M., Firshein W.;
RT "Identification of a potential membrane-targeting region of the replication
RT initiator protein (TrfA) of broad-host-range plasmid RK2.";
RL Plasmid 43:214-222(2000).
RN [7]
RP INTERACTION WITH HDA.
RC STRAIN=B / BL21-DE3; PLASMID=IncP-alpha RK2;
RX PubMed=12618445; DOI=10.1128/jb.185.6.1817-1824.2003;
RA Kim P.D., Banack T., Lerman D.M., Tracy J.C., Camara J.E., Crooke E.,
RA Oliver D., Firshein W.;
RT "Identification of a novel membrane-associated gene product that suppresses
RT toxicity of a TrfA peptide from plasmid RK2 and its relationship to the
RT DnaA host initiation protein.";
RL J. Bacteriol. 185:1817-1824(2003).
CC -!- FUNCTION: Required for initiation of plasmid DNA replication, along
CC with host-derived DnaA and other host proteins. Both forms of the
CC protein are capable of initiating plasmid replication in a number of
CC Gram-negative bacteria. Binds to 8 17-base pair repeat sequences
CC (iterons) in the RK2 minimal replication origin (oriV), opening the
CC origin of replication. oriV opening does not absolutely require the
CC presence of nucleotides; formation of open complex is somewhat enhanced
CC by ATP or ATP gamma S, while DnaA or HU is required for full opening.
CC -!- FUNCTION: Also involved in plasmid copy number control, promoting
CC intermolecular coupling of protein bound iterons at oriV, which
CC inhibits replication initiation.
CC -!- SUBUNIT: Forms a dimer in solution, binds DNA as a monomer. Both
CC mononer and dimer of the short form interact with Hda (Dp).
CC {ECO:0000269|PubMed:8636140}.
CC -!- SUBCELLULAR LOCATION: Cell inner membrane {ECO:0000269|PubMed:10692384,
CC ECO:0000269|PubMed:10783300}; Peripheral membrane protein
CC {ECO:0000269|PubMed:10692384, ECO:0000269|PubMed:10783300}.
CC Note=Probably the site where plasmid replication is initiated.
CC -!- ALTERNATIVE PRODUCTS:
CC Event=Alternative initiation; Named isoforms=2;
CC Name=TrfA;
CC IsoId=P07676-1; Sequence=Displayed;
CC Name=TrfA*;
CC IsoId=P07676-2; Sequence=VSP_018862;
CC -!- MISCELLANEOUS: This broad-host-range plasmid is capable of replicating
CC in a number of Gram-negative bacteria.
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DR EMBL; X00713; CAA25306.1; -; Genomic_DNA.
DR PIR; S08595; S08595.
DR RefSeq; WP_001082279.1; NZ_NJTR01000057.1.
DR AlphaFoldDB; P07676; -.
DR SMR; P07676; -.
DR GO; GO:0005886; C:plasma membrane; IEA:UniProtKB-SubCell.
DR GO; GO:0003677; F:DNA binding; IEA:UniProtKB-KW.
DR GO; GO:0006260; P:DNA replication; IEA:UniProtKB-KW.
DR GO; GO:0006276; P:plasmid maintenance; IEA:UniProtKB-KW.
DR InterPro; IPR010751; TrfA.
DR Pfam; PF07042; TrfA; 1.
PE 1: Evidence at protein level;
KW Alternative initiation; Cell inner membrane; Cell membrane;
KW DNA replication; DNA-binding; Membrane; Plasmid; Plasmid copy control.
FT CHAIN 1..382
FT /note="Plasmid replication initiator protein TrfA"
FT /id="PRO_0000024513"
FT DNA_BIND 246..265
FT /note="H-T-H motif"
FT /evidence="ECO:0000305"
FT REGION 1..163
FT /note="Toxic in E.coli strain K12 / DH5-alpha; may be
FT membrane-associated"
FT REGION 286..297
FT /note="Hydrophobic region (HR); required for membrane
FT association"
FT VAR_SEQ 1..97
FT /note="Missing (in isoform TrfA*)"
FT /evidence="ECO:0000305"
FT /id="VSP_018862"
FT MUTAGEN 254
FT /note="G->D: 16-fold increased plasmid copy number.
FT Uncontrolled replication, in the active monomer form,
FT increased DNA-binding, loss of intermolecular coupling;
FT when associated with L-267."
FT /evidence="ECO:0000269|PubMed:8622975,
FT ECO:0000269|PubMed:9242693"
FT MUTAGEN 267
FT /note="S->L: 23-fold increased plasmid copy number.
FT Uncontrolled in the active monomer form, increased DNA-
FT binding, loss of intermolecular coupling; when associated
FT with D-254."
FT /evidence="ECO:0000269|PubMed:8622975,
FT ECO:0000269|PubMed:9242693"
FT MUTAGEN 286..297
FT /note="VLIDEEIVVLFA->SR: Loss of membrane association, no
FT plasmid replication, less protein produced."
FT /evidence="ECO:0000269|PubMed:10783300"
FT MUTAGEN 289..291
FT /note="DEE->AAA: Decreased membrane association, impaired
FT plasmid replication."
FT /evidence="ECO:0000269|PubMed:10783300"
FT MUTAGEN 289..291
FT /note="DEE->NQQ: No change in membrane association,
FT impaired plasmid replication."
FT /evidence="ECO:0000269|PubMed:10783300"
FT MUTAGEN 293..294
FT /note="VV->TT: Loss of membrane association, no plasmid
FT replication, less protein produced."
FT /evidence="ECO:0000269|PubMed:10783300"
FT MUTAGEN 296
FT /note="F->A: Decreased membrane association, impaired
FT plasmid replication."
FT /evidence="ECO:0000269|PubMed:10783300"
SQ SEQUENCE 382 AA; 43766 MW; 944FFD1FA0A93869 CRC64;
MNRTFDRKAY RQELIDAGFS AEDAETIASR TVMRAPRETF QSVGSMVQQA TAKIERDSVQ
LAPPALPAPS AAVERSRRLE QEAAGLAKSM TIDTRGTMTT KKRKTAGEDL AKQVSEAKQA
ALLKHTKQQI KEMQLSLFDI APWPDTMRAM PNDTARSALF TTRNKKIPRE ALQNKVIFHV
NKDVKITYTG VELRADDDEL VWQQVLEYAK RTPIGEPITF TFYELCQDLG WSINGRYYTK
AEECLSRLQA TAMGFTSDRV GHLESVSLLH RFRVLDRGKK TSRCQVLIDE EIVVLFAGDH
YTKFIWEKYR KLSPTARRMF DYFSSHREPY PLKLETFRLM CGSDSTRVKK WREQVGEACE
ELRGSGLVEH AWVNDDLVHC KR
