Detailed information of oriT
oriT
The information of the oriT region
| oriTDB ID | 100008 |
| Name | oriT_R388  |
| Organism | Escherichia coli |
| Sequence Completeness | intact |
| NCBI accession of oriT (coordinates [strand]) | NC_028464 (15948..16349 [-], 402 nt) |
| oriT length | 402 nt |
| IRs (inverted repeats) | IR1: 135..141, 148..154 (TGTAGTG..CACTACA) IR2: 177..184, 192..199 (AAGTCATT..AATGACTT) IR3: 203..208, 213..218 (CGCACC..GGTGCG) IR4: 317..326, 330..339 (CCCAATGCGC..GCGCATTGGG) |
| Location of nic site | 226..227 |
| Conserved sequence flanking the nic site |
TGTCT|ATAGC |
| Note | (1) The relaxase TrwC_R388 is transported by the T4SS of plasmid R388, and it can remain active in the recipient cell [PMID:16260740]. (2) The T4CP TrwB_R388 ATPase activity is stimulated by ssDNA, dsDNA and the C-terminal Region of auxiliary protein TrwA_R388 [PMID:17599913]. (3) In oriT_R388, the relaxase TrwC_R388 specific binding residues exclusively in the region 5’ to nic site. [PMID:14625590]. (4) GTAGTG in oriT_R388 is the specific binding sequence of the auxiliary protein TrwA_R388 [PMID:15450172]. (5) The auxiliary protein TrwA_R388 binds specifically to two regions (region A and region B) with different affinities: Region A (high affinity) : 71-108 bp and Region B (low affinity) : 125-165 bp [PMID:9236121]. (6) The auxiliary protein TrwA_R388 perform two biochemical activities: binding to oriT resulted in transcriptional repression of the trwABC operon and enhanced the relaxation activity of relaxase TrwC_R388 in vitro [PMID:9236121]. (7) The relaxase TrwC_R388 contains two active-site tyrosyl residues, Y18 and Y26, that act sequentially at the origin of transfer during plasmid R388 conjugation [PMID:10653694]. (8) Mutation Y18F, but not mutation Y26F, abolished nic-cleavage of a supercoiled DNA containing the oriT_R388 [PMID:10653694]. (9) The auxiliary protein IHF_R388 have an inhibitory role in the nic-cleavage reaction in vivo [PMID:10209739]. |
oriT sequence
Download Length: 402 nt
CCGCCTCGTCCTCCAAAAGTGCCTGCTTTTCCGGGCTTAGCCGTACTTGGATGGGGTCGCCTAGTGCCATGTCCTCTCCCGTAGTGTTACTGTAGTGGTTCAATCCTAGCATTTACAAGGGGTTGCGGCAATATTGTAGTGGCATAACACTACACAGGTTTTCGTCCTTGGCGTGGAAGTCATTGTAAATCAATGACTTACGCGCACCGAAAGGTGCGTATTGTCTATAGCCCAGATTTAAGGATACCAACCCGGCTTTTAAGGACGGAAACCATGCGATAACGCCAGCGTGACCCTAAAGAGGGTCAAAACTGCTCCCAATGCGCTATGCGCATTGGGTTATCGTGCAGCAATGATGCAACTATAATGCTATGATGGTGCTACAATGATGCAGAAAATGAG
Visualization of oriT structure (The oriT was characterized experimentally)
Base pairs in green indicate regions allow point mutation.
Base pairs in yellow indicate regions allow partial point mutation.
Base pairs in red indicate regions do not allow point mutation.
(Reference: PMID:25123661)
oriT secondary structure
Predicted by RNAfold.
Download structure file
Reference
[1] Revilla C et al. (2008) Different pathways to acquiring resistance genes illustrated by the recent evolution of IncW plasmids. Antimicrob Agents Chemother. 52(4):1472-80. [PMID:18268088]
[2] Paterson ES et al. (1997) Localization of the nic site of IncN conjugative plasmid pCU1 through formation of a hybrid oriT. J Bacteriol. 179(18):5768-76. [PMID:9294433]
[3] Llosa M et al. (1995) Nicking activity of TrwC directed against the origin of transfer of the IncW plasmid R388. J Mol Biol. 246(1):54-62. [PMID:7853404]
[4] Llosa M et al. (1991) Structural and functional analysis of the origin of conjugal transfer of the broad-host-range IncW plasmid R388 and comparison with the related IncN plasmid R46. Mol Gen Genet. 226(3):473-83. [PMID:2038309]
Relaxosome
This oriT is a component of a relaxosome.
| Relaxosome name | RelaxosomeR388  |
| oriT | oriT_R388 |
| Relaxase | TrwC_R388 (MOBF) |
| Auxiliary protein | TrwA_R388 |
Reference
[1] de la Cruz F et al. (2010) Conjugative DNA metabolism in Gram-negative bacteria. FEMS Microbiol Rev. 34(1):18-40. [PMID:19919603]
[2] Revilla C et al. (2008) Different pathways to acquiring resistance genes illustrated by the recent evolution of IncW plasmids. Antimicrob Agents Chemother. 52(4):1472-80. [PMID:18268088]
[3] Fernández-López R et al. (2006) Dynamics of the IncW genetic backbone imply general trends in conjugative plasmid evolution. FEMS Microbiol Rev. 30(6):942-66. [PMID:17026718]
[4] Paterson ES et al. (1997) Localization of the nic site of IncN conjugative plasmid pCU1 through formation of a hybrid oriT. J Bacteriol. 179(18):5768-76. [PMID:9294433]
[5] Llosa M et al. (1995) Nicking activity of TrwC directed against the origin of transfer of the IncW plasmid R388. J Mol Biol. 246(1):54-62. [PMID:7853404]
[6] Llosa M et al. (1994) Genetic organization of the conjugal DNA processing region of the IncW plasmid R388. J Mol Biol. 235(2):448-64. [PMID:8289274]
[7] Llosa M et al. (1991) Structural and functional analysis of the origin of conjugal transfer of the broad-host-range IncW plasmid R388 and comparison with the related IncN plasmid R46. Mol Gen Genet. 226(3):473-83. [PMID:2038309]
Relaxase
| ID | 7 | GenBank | FAA00039 |
| Name | TrwC_R388 |
UniProt ID | Q2NKF7 |
| Length | 966 a.a. | PDB ID | |
| Note | (1) The relaxase TrwC_R388 is transported by the T4SS of plasmid R388, and it can remain active in the recipient cell [PMID:16260740]. (2) In oriT_R388, the relaxase TrwC_R388 specific binding residues exclusively in the region 5' to nic site. [PMID:14625590]. (3) The relaxase TrwC_R388 contains two active-site tyrosyl residues, Y18 and Y26, that act sequentially at the origin of transfer during plasmid R388 conjugation [PMID:10653694]. |
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Relaxase protein sequence
Download Length: 966 a.a. Molecular weight: 107676.06 Da Isoelectric Point: 9.9900
MLSHMVLTRQDIGRAASYYEDGADDYYAKDGDASEWQGKGAEELGLSGEVDSKRFRELLAGNIGEGHRIM
RSATRQDSKERIGLDLTFSAPKSVSLQALVAGDAEIIKAHDRAVARTLEQAEARAQARQKIQGKTRIETT
GNLVIGKFRHETSRERDPQLHTHAVILNMTKRSDGQWRALKNDEIVKRTRYLGAVYNAELAHELQKLGYQ
LRYGKDGNFDLAHIDRQQIEGFSKRTEQIAEWYAARGLDPNSVSLEQKQAAKVLSRAKKTSVDREALRAE
WQATAKELGIDFSRREWSGREKGGSEKQAHSFMPSDEAAKRAVRYAINHLTERQSVMDERELVDTAMKHA
VGAARLEDIQKELLRQTETGYLIREAPRYRPGGQTGPTDEPGKTRAEWVAELAAKGMKQGAARERVDNAI
KTGGLVPIEPRYTTQTALEREKRILQIERDGRGAVAPVIAAEAARERLASTNLNQGQREAAELIVSAANR
VVGVQGFAGTGKSHMLDTAKQMIEGEGYHVRALAAYGSQVKALRELNVEANTLASFLRAKDKNIDSRTVL
VIDEAGVVPTRLMEQTLKLAEKAGARVVLMGDTAQTKAIEAGRPFDQLQAAGMQTAHMREIQRQKNPELK
IAVELAAAGKASSSLERIKDVTEIKNHHERRAAVAEAYIALKPDERDRTLIVSGTNEARREINQIVREGL
GTAGKGIEFDTLVRVDTTQAERRHSKNYQVGHVIQPERDYAKTGLQRGELYRVVETGPGNRLTVIGEHDG
QRIQFSPMTHTKISVYQPERAELAVGDTIRITRNDKHLDLANGDRMKVVAVEDRKVTVTDGKRNVELPTD
KPLHVDHAYATTVHSSQGLTSDRVLIDAHAESRTTRKDVYYVAISRARFEARVFTNDRGKLPAAIARENI
KSAAHDLARDRGGRSAAAERQREQQRERERNRQTQQPAHDRQKAAREAERGMEAGR
Protein domains
Predicted by InterproScan.
Protein structure
| Source | ID | Structure |
|---|---|---|
| AlphaFold DB | Q2NKF7 |
Reference
[1] de la Cruz F et al. (2010) Conjugative DNA metabolism in Gram-negative bacteria. FEMS Microbiol Rev. 34(1):18-40. [PMID:19919603]
[2] Revilla C et al. (2008) Different pathways to acquiring resistance genes illustrated by the recent evolution of IncW plasmids. Antimicrob Agents Chemother. 52(4):1472-80. [PMID:18268088]
[3] Fernández-López R et al. (2006) Dynamics of the IncW genetic backbone imply general trends in conjugative plasmid evolution. FEMS Microbiol Rev. 30(6):942-66. [PMID:17026718]
[4] Llosa M et al. (1995) Nicking activity of TrwC directed against the origin of transfer of the IncW plasmid R388. J Mol Biol. 246(1):54-62. [PMID:7853404]
[5] Llosa M et al. (1994) Genetic organization of the conjugal DNA processing region of the IncW plasmid R388. J Mol Biol. 235(2):448-64. [PMID:8289274]
Auxiliary protein
| ID | 13 | GenBank | YP_009182121 |
| Name | TrwA_R388 |
UniProt ID | Q04229 |
| Length | 121 a.a. | PDB ID | _ |
| Note | (1) The T4CP TrwB_R388 ATPase activity is stimulated by ssDNA, dsDNA and the C-terminal Region of auxiliary protein TrwA_R388 [PMID:17599913]. (2) GTAGTG in oriT_R388 is the specific binding sequence of the auxiliary protein TrwA_R388 [PMID:15450172]. (3) The auxiliary protein TrwA_R388 binds specifically to two regions (region A and region B) with different affinities: Region A (high affinity) : 71-108 bp and Region B (low affinity) : 125-165 bp [PMID:9236121]. (4) The auxiliary protein TrwA_R388 perform two biochemical activities: binding to oriT resulted in transcriptional repression of the trwABC operon and enhanced the relaxation activity of relaxase TrwC_R388 in vitro [PMID:9236121]. |
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Auxiliary protein sequence
Download Length: 121 a.a. Molecular weight: 13400.32 Da Isoelectric Point: 4.8464
MALGDPIQVRLSPEKQALLEDEAARKGKRLATYLRELLESENDLQGELAALRREVVSLHHVIEDLADTGL
RSDQSGPGQNAVQIETLLLLRAIAGPERMKPVKGELKRLGIEVWTPEGKED
Protein domains
No domain identified.
Protein structure
| Source | ID | Structure |
|---|---|---|
| AlphaFold DB | Q04229 |
Reference
[1] de la Cruz F et al. (2010) Conjugative DNA metabolism in Gram-negative bacteria. FEMS Microbiol Rev. 34(1):18-40. [PMID:19919603]
[2] Revilla C et al. (2008) Different pathways to acquiring resistance genes illustrated by the recent evolution of IncW plasmids. Antimicrob Agents Chemother. 52(4):1472-80. [PMID:18268088]
[3] Fernández-López R et al. (2006) Dynamics of the IncW genetic backbone imply general trends in conjugative plasmid evolution. FEMS Microbiol Rev. 30(6):942-66. [PMID:17026718]
[4] Llosa M et al. (1994) Genetic organization of the conjugal DNA processing region of the IncW plasmid R388. J Mol Biol. 235(2):448-64. [PMID:8289274]
T4CP
| ID | 7 | GenBank | FAA00040 |
| Name | TrwB_R388 |
UniProt ID | Q04230 |
| Length | 507 a.a. | PDB ID | 1E9R, 1E9S, 1GKI, 1GL6, 1GL7 |
| Note | The T4CP TrwB_R388 ATPase activity is stimulated by ssDNA, dsDNA and the C-terminal Region of auxiliary protein TrwA_R388 [PMID:17599913]. | ||
T4CP protein sequence
Download Length: 507 a.a. Molecular weight: 56272.95 Da Isoelectric Point: 9.8647
MHPDDQRKVSAGIVIVLPLIFWITAVQKTEVLGSPKLLALWELMKLTPQKPILLLSALGGLAVGVLFVWL
LNSVGQGEFGGAPFKRFLRGTRIVSGGKLKRMTREKAKQVTVAGVPMPRDAEPRHLLVNGATGTGKSVLL
RELAYTGLLRGDRMVIVDPNGDMLSKFGRDKDIILNPYDQRTKGWSFFNEIRNDYDWQRYALSVVPRGKT
DEAEEWASYGRLLLRETAKKLALIGTPSMRELFHWTTIATFDDLRGFLEGTLAESLFAGSNEASKALTSA
RFVLSDKLPEHVTMPDGDFSIRSWLEDPNGGNLFITWREDMGPALRPLISAWVDVVCTSILSLPEEPKRR
LWLFIDELASLEKLASLADALTKGRKAGLRVVAGLQSTSQLDDVYGVKEAQTLRASFRSLVVLGGSRTDP
KTNEDMSLSLGEHEVERDRYSKNTGKHHSTGRALERVRERVVMPAEIANLPDLTAYVGFAGNRPIAKVPL
EIKQFANRQPAFVEGTI
Protein domains
Predicted by InterproScan.
Protein structure
| Source | ID | Structure |
|---|---|---|
| PDB | 1E9R | |
| PDB | 1E9S | |
| PDB | 1GKI | |
| PDB | 1GL6 | |
| PDB | 1GL7 | |
| AlphaFold DB | Q04230 |
Reference
[1] de la Cruz F et al. (2010) Conjugative DNA metabolism in Gram-negative bacteria. FEMS Microbiol Rev. 34(1):18-40. [PMID:19919603]
[2] Fernández-López R et al. (2006) Dynamics of the IncW genetic backbone imply general trends in conjugative plasmid evolution. FEMS Microbiol Rev. 30(6):942-66. [PMID:17026718]
[3] Llosa M et al. (1994) Genetic organization of the conjugal DNA processing region of the IncW plasmid R388. J Mol Biol. 235(2):448-64. [PMID:8289274]
[4] Cabezón E et al. (1994) Requirements for mobilization of plasmids RSF1010 and ColE1 by the IncW plasmid R388: trwB and RP4 traG are interchangeable. J Bacteriol. 176(14):4455-8. [PMID:8021231]
T4SS
T4SS were predicted by using oriTfinder2.
Region 1: 1020..15650
| Locus tag | Coordinates | Strand | Size (bp) | Protein ID | Product | Description |
|---|---|---|---|---|---|---|
| HXF36_RS00005 | 1..288 | - | 288 | WP_012414174 | H-NS histone family protein | - |
| HXF36_RS00010 | 437..1018 | + | 582 | WP_012196419 | hypothetical protein | - |
| HXF36_RS00015 | 1020..1616 | + | 597 | WP_032491146 | lytic transglycosylase domain-containing protein | virB1 |
| HXF36_RS00020 | 1841..2137 | + | 297 | WP_032491147 | KorA family transcriptional regulator | - |
| HXF36_RS00025 | 2103..2408 | + | 306 | WP_031942907 | VirB2 family type IV secretion system major pilin TrwL | virB2 |
| HXF36_RS00030 | 2421..2735 | + | 315 | WP_012443546 | VirB3 family type IV secretion system protein | virB3 |
| HXF36_RS00035 | 2738..5209 | + | 2472 | WP_250693943 | VirB4 family type IV secretion/conjugal transfer ATPase | virb4 |
| HXF36_RS00040 | 5206..5886 | + | 681 | WP_032490329 | type IV secretion system protein | virB5 |
| HXF36_RS00045 | 5883..6113 | + | 231 | WP_012196425 | EexN family lipoprotein | - |
| HXF36_RS00050 | 6127..7155 | + | 1029 | WP_032490330 | type IV secretion system protein | virB6 |
| HXF36_RS00055 | 7409..8104 | + | 696 | WP_032490331 | VirB8/TrbF family protein | virB8 |
| HXF36_RS00060 | 8115..8915 | + | 801 | WP_032490332 | TrbG/VirB9 family P-type conjugative transfer protein | virB9 |
| HXF36_RS00065 | 8915..10102 | + | 1188 | WP_032490333 | type IV secretion system protein VirB10 | virB10 |
| HXF36_RS00070 | 10068..11144 | + | 1077 | WP_012196431 | P-type DNA transfer ATPase VirB11 | virB11 |
| HXF36_RS00075 | 11227..14115 | - | 2889 | WP_250693944 | MobF family relaxase | - |
| HXF36_RS00080 | 14127..15650 | - | 1524 | WP_012196433 | type IV secretion system DNA-binding domain-containing protein | virb4 |
| HXF36_RS00085 | 15652..16017 | - | 366 | WP_012196434 | hypothetical protein | - |
| HXF36_RS00090 | 16471..16896 | + | 426 | WP_032491149 | hypothetical protein | - |
| HXF36_RS00095 | 16893..17588 | + | 696 | WP_012196436 | StbB family protein | - |
| HXF36_RS00100 | 17602..17988 | + | 387 | WP_032491150 | hypothetical protein | - |
| HXF36_RS00105 | 18142..18393 | - | 252 | WP_012196438 | hypothetical protein | - |
| HXF36_RS00110 | 18541..19194 | - | 654 | WP_012196439 | hypothetical protein | - |
| HXF36_RS00115 | 19210..19554 | - | 345 | WP_012196440 | hypothetical protein | - |
| HXF36_RS00120 | 20247..20570 | - | 324 | WP_012196442 | TrfB-related DNA-binding protein | - |
Host bacterium
| ID | 7 | GenBank | NC_028464 |
| Plasmid name | R388 | Incompatibility group | IncW |
| Plasmid size | 33913 bp | Coordinate of oriT [Strand] | 15948..16349 [-] |
| Host baterium | Escherichia coli |
Cargo genes
| Drug resistance gene | dfrB2, qacE, sul1 |
| Virulence gene | trwM, trwK, trwG, trwF, trwD |
| Metal resistance gene | _ |
| Degradation gene | _ |
| Symbiosis gene | - |
| Anti-CRISPR | - |