TTUA_THET2
ID TTUA_THET2 Reviewed; 321 AA.
AC Q72LF3;
DT 22-NOV-2017, integrated into UniProtKB/Swiss-Prot.
DT 05-JUL-2004, sequence version 1.
DT 03-AUG-2022, entry version 102.
DE RecName: Full=tRNA-5-methyluridine(54) 2-sulfurtransferase {ECO:0000305|PubMed:16547008};
DE EC=2.8.1.15 {ECO:0000269|PubMed:28439027, ECO:0000305|PubMed:16547008};
DE AltName: Full=2-thiouridine synthetase TtuA {ECO:0000303|PubMed:28439027};
DE AltName: Full=tRNA two-thiouridine-synthesizing protein A {ECO:0000303|PubMed:16547008};
GN Name=ttuA {ECO:0000303|PubMed:16547008};
GN OrderedLocusNames=TT_C0106 {ECO:0000312|EMBL:AAS80454.1};
OS Thermus thermophilus (strain ATCC BAA-163 / DSM 7039 / HB27).
OC Bacteria; Deinococcus-Thermus; Deinococci; Thermales; Thermaceae; Thermus.
OX NCBI_TaxID=262724;
RN [1]
RP NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA].
RC STRAIN=ATCC BAA-163 / DSM 7039 / HB27;
RX PubMed=15064768; DOI=10.1038/nbt956;
RA Henne A., Brueggemann H., Raasch C., Wiezer A., Hartsch T., Liesegang H.,
RA Johann A., Lienard T., Gohl O., Martinez-Arias R., Jacobi C.,
RA Starkuviene V., Schlenczeck S., Dencker S., Huber R., Klenk H.-P.,
RA Kramer W., Merkl R., Gottschalk G., Fritz H.-J.;
RT "The genome sequence of the extreme thermophile Thermus thermophilus.";
RL Nat. Biotechnol. 22:547-553(2004).
RN [2]
RP IDENTIFICATION BY MASS SPECTROMETRY, FUNCTION, CATALYTIC ACTIVITY,
RP TRNA-BINDING, DISRUPTION PHENOTYPE, PATHWAY, AND SUBUNIT.
RC STRAIN=ATCC BAA-163 / DSM 7039 / HB27;
RX PubMed=16547008; DOI=10.1074/jbc.m511675200;
RA Shigi N., Sakaguchi Y., Suzuki T., Watanabe K.;
RT "Identification of two tRNA thiolation genes required for cell growth at
RT extremely high temperatures.";
RL J. Biol. Chem. 281:14296-14306(2006).
RN [3]
RP CONJUGATION TO TTUB, AND ACTIVITY REGULATION.
RC STRAIN=ATCC BAA-163 / DSM 7039 / HB27;
RX PubMed=22467871; DOI=10.1074/jbc.m112.359844;
RA Shigi N.;
RT "Posttranslational modification of cellular proteins by a ubiquitin-like
RT protein in bacteria.";
RL J. Biol. Chem. 287:17568-17577(2012).
RN [4]
RP MUTAGENESIS OF LYS-58; ASP-59; CYS-130; CYS-133; HIS-157; ASN-158; ASP-161;
RP GLU-203 AND CYS-222.
RC STRAIN=ATCC BAA-163 / DSM 7039 / HB27;
RX PubMed=23444054; DOI=10.1002/prot.24273;
RA Nakagawa H., Kuratani M., Goto-Ito S., Ito T., Katsura K., Terada T.,
RA Shirouzu M., Sekine S., Shigi N., Yokoyama S.;
RT "Crystallographic and mutational studies on the tRNA thiouridine synthetase
RT TtuA.";
RL Proteins 81:1232-1244(2013).
RN [5] {ECO:0007744|PDB:5B4E, ECO:0007744|PDB:5B4F, ECO:0007744|PDB:5GHA}
RP X-RAY CRYSTALLOGRAPHY (2.50 ANGSTROMS) IN COMPLEXES WITH ATP ANALOG;
RP IRON-SULFUR (4FE-4S); ZINC AND TTUB PROTEIN, FUNCTION, CATALYTIC ACTIVITY,
RP COFACTOR, SUBUNIT, REACTION MECHANISM, AND MUTAGENESIS OF CYS-130; CYS-133
RP AND CYS-222.
RX PubMed=28439027; DOI=10.1073/pnas.1615585114;
RA Chen M., Asai S., Narai S., Nambu S., Omura N., Sakaguchi Y., Suzuki T.,
RA Ikeda-Saito M., Watanabe K., Yao M., Shigi N., Tanaka Y.;
RT "Biochemical and structural characterization of oxygen-sensitive 2-
RT thiouridine synthesis catalyzed by an iron-sulfur protein TtuA.";
RL Proc. Natl. Acad. Sci. U.S.A. 114:4954-4959(2017).
CC -!- FUNCTION: Catalyzes the ATP-dependent 2-thiolation of 5-methyluridine
CC residue at position 54 in the T loop of tRNAs, leading to 5-methyl-2-
CC thiouridine (m(5)s(2)U or s(2)T) (PubMed:16547008, PubMed:28439027).
CC This modification allows thermal stabilization of tRNAs in thermophilic
CC microorganisms, and is required for cell growth at high temperatures
CC (PubMed:16547008). TtuA transfers the S atom from the thiocarboxylated
CC C-terminus of TtuB to tRNA (PubMed:28439027).
CC {ECO:0000269|PubMed:16547008, ECO:0000269|PubMed:28439027}.
CC -!- CATALYTIC ACTIVITY:
CC Reaction=5-methyluridine(54) in tRNA + [TtuB sulfur-carrier protein]-
CC Gly-NH-CH2-C(O)SH + ATP + H2O = 5-methyl-2-thiouridine(54) in tRNA +
CC [TtuB sulfur-carrier protein]-C-terminal Gly-Gly + AMP + diphosphate
CC + 2 H(+); Xref=Rhea:RHEA:52720, Rhea:RHEA-COMP:10167, Rhea:RHEA-
CC COMP:13342, Rhea:RHEA-COMP:13343, Rhea:RHEA-COMP:13344,
CC ChEBI:CHEBI:15377, ChEBI:CHEBI:15378, ChEBI:CHEBI:30616,
CC ChEBI:CHEBI:33019, ChEBI:CHEBI:74447, ChEBI:CHEBI:90619,
CC ChEBI:CHEBI:90778, ChEBI:CHEBI:136799, ChEBI:CHEBI:456215;
CC EC=2.8.1.15; Evidence={ECO:0000269|PubMed:28439027,
CC ECO:0000305|PubMed:16547008};
CC -!- COFACTOR:
CC Name=[4Fe-4S] cluster; Xref=ChEBI:CHEBI:49883;
CC Evidence={ECO:0000269|PubMed:28439027};
CC Note=Binds 1 [4Fe-4S] cluster per subunit. The cluster is chelated by
CC three Cys residues, the Fe with a free coordination site of the Fe-S
CC cluster may contribute to an enzymatic role.
CC {ECO:0000269|PubMed:28439027};
CC -!- COFACTOR:
CC Name=Mg(2+); Xref=ChEBI:CHEBI:18420;
CC Evidence={ECO:0000250|UniProtKB:Q9WY40};
CC -!- ACTIVITY REGULATION: Enzymatic activity may be regulated by TtuB
CC conjugation. {ECO:0000305|PubMed:22467871}.
CC -!- PATHWAY: tRNA modification. {ECO:0000269|PubMed:16547008}.
CC -!- SUBUNIT: Homodimer (PubMed:28439027). Is able to form a heterocomplex
CC with TtuB (PubMed:16547008, PubMed:28439027).
CC {ECO:0000269|PubMed:16547008, ECO:0000269|PubMed:28439027}.
CC -!- PTM: Conjugated to TtuB via covalent linkages involving Lys-137, Lys-
CC 226 and Lys-229. {ECO:0000269|PubMed:22467871}.
CC -!- DISRUPTION PHENOTYPE: Disruption of this gene completely abolishes the
CC presence of m(5)s(2)U in tRNAs, although its precursor, 5-methyluridine
CC (ribothymidine) is present; other nucleoside modifications remain
CC unchanged. These deletion mutants exhibit a temperature-sensitive
CC phenotype, they are unable to grow above 80 degrees Celsius.
CC {ECO:0000269|PubMed:16547008}.
CC -!- MISCELLANEOUS: The thiolation reaction consists of two steps: a first
CC activation step by ATP to form an adenylated intermediate of the target
CC base of tRNA, and a second nucleophilic substitution step of the sulfur
CC (S) atom supplied by thiocarboxylated TtuB for the adenyl group.
CC However, it is not clear if the S-transfer mechanism is direct or
CC indirect. {ECO:0000305|PubMed:28439027}.
CC -!- SIMILARITY: Belongs to the TtcA family. TtuA subfamily. {ECO:0000305}.
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DR EMBL; AE017221; AAS80454.1; -; Genomic_DNA.
DR RefSeq; WP_011172563.1; NC_005835.1.
DR PDB; 5B4E; X-ray; 2.70 A; A=1-321.
DR PDB; 5B4F; X-ray; 2.75 A; A=1-321.
DR PDB; 5GHA; X-ray; 2.50 A; A/B/C/D=1-321.
DR PDBsum; 5B4E; -.
DR PDBsum; 5B4F; -.
DR PDBsum; 5GHA; -.
DR AlphaFoldDB; Q72LF3; -.
DR SMR; Q72LF3; -.
DR STRING; 262724.TT_C0106; -.
DR PRIDE; Q72LF3; -.
DR EnsemblBacteria; AAS80454; AAS80454; TT_C0106.
DR KEGG; tth:TT_C0106; -.
DR eggNOG; COG0037; Bacteria.
DR HOGENOM; CLU_026481_1_1_0; -.
DR OMA; MNLDQKQ; -.
DR OrthoDB; 1051352at2; -.
DR BioCyc; MetaCyc:MON-20129; -.
DR BRENDA; 2.8.1.15; 2305.
DR Proteomes; UP000000592; Chromosome.
DR GO; GO:0051539; F:4 iron, 4 sulfur cluster binding; IEA:UniProtKB-KW.
DR GO; GO:0005524; F:ATP binding; IEA:UniProtKB-KW.
DR GO; GO:0046872; F:metal ion binding; IEA:UniProtKB-KW.
DR GO; GO:0016740; F:transferase activity; IEA:UniProtKB-KW.
DR GO; GO:0000049; F:tRNA binding; IEA:UniProtKB-KW.
DR GO; GO:0034227; P:tRNA thio-modification; IEA:InterPro.
DR GO; GO:0002098; P:tRNA wobble uridine modification; IEA:InterPro.
DR Gene3D; 3.40.50.620; -; 1.
DR InterPro; IPR000541; Ncs6/Tuc1/Ctu1.
DR InterPro; IPR014729; Rossmann-like_a/b/a_fold.
DR InterPro; IPR011063; TilS/TtcA_N.
DR InterPro; IPR035107; tRNA_thiolation_TtcA_Ctu1.
DR Pfam; PF01171; ATP_bind_3; 1.
DR PIRSF; PIRSF004976; ATPase_YdaO; 1.
DR TIGRFAMs; TIGR00269; TIGR00269; 1.
PE 1: Evidence at protein level;
KW 3D-structure; 4Fe-4S; ATP-binding; Iron; Iron-sulfur; Isopeptide bond;
KW Magnesium; Metal-binding; Nucleotide-binding; RNA-binding; Transferase;
KW tRNA processing; tRNA-binding; Ubl conjugation; Zinc.
FT CHAIN 1..321
FT /note="tRNA-5-methyluridine(54) 2-sulfurtransferase"
FT /id="PRO_0000442363"
FT BINDING 3
FT /ligand="Zn(2+)"
FT /ligand_id="ChEBI:CHEBI:29105"
FT /ligand_label="1"
FT /evidence="ECO:0000269|PubMed:28439027,
FT ECO:0007744|PDB:5B4E, ECO:0007744|PDB:5B4F,
FT ECO:0007744|PDB:5GHA"
FT BINDING 6
FT /ligand="Zn(2+)"
FT /ligand_id="ChEBI:CHEBI:29105"
FT /ligand_label="1"
FT /evidence="ECO:0000269|PubMed:28439027,
FT ECO:0007744|PDB:5B4E, ECO:0007744|PDB:5B4F,
FT ECO:0007744|PDB:5GHA"
FT BINDING 22
FT /ligand="Zn(2+)"
FT /ligand_id="ChEBI:CHEBI:29105"
FT /ligand_label="1"
FT /evidence="ECO:0000269|PubMed:28439027,
FT ECO:0007744|PDB:5B4E, ECO:0007744|PDB:5B4F,
FT ECO:0007744|PDB:5GHA"
FT BINDING 25
FT /ligand="Zn(2+)"
FT /ligand_id="ChEBI:CHEBI:29105"
FT /ligand_label="1"
FT /evidence="ECO:0000269|PubMed:28439027,
FT ECO:0007744|PDB:5B4E, ECO:0007744|PDB:5B4F,
FT ECO:0007744|PDB:5GHA"
FT BINDING 53..55
FT /ligand="ATP"
FT /ligand_id="ChEBI:CHEBI:30616"
FT /evidence="ECO:0000269|PubMed:28439027,
FT ECO:0007744|PDB:5B4E"
FT BINDING 59
FT /ligand="ATP"
FT /ligand_id="ChEBI:CHEBI:30616"
FT /evidence="ECO:0000269|PubMed:28439027,
FT ECO:0007744|PDB:5B4E"
FT BINDING 79
FT /ligand="ATP"
FT /ligand_id="ChEBI:CHEBI:30616"
FT /evidence="ECO:0000269|PubMed:28439027,
FT ECO:0007744|PDB:5B4E"
FT BINDING 130
FT /ligand="[4Fe-4S] cluster"
FT /ligand_id="ChEBI:CHEBI:49883"
FT /evidence="ECO:0000269|PubMed:28439027,
FT ECO:0007744|PDB:5B4E, ECO:0007744|PDB:5B4F"
FT BINDING 133
FT /ligand="[4Fe-4S] cluster"
FT /ligand_id="ChEBI:CHEBI:49883"
FT /evidence="ECO:0000269|PubMed:28439027,
FT ECO:0007744|PDB:5B4E, ECO:0007744|PDB:5B4F"
FT BINDING 156
FT /ligand="ATP"
FT /ligand_id="ChEBI:CHEBI:30616"
FT /evidence="ECO:0000269|PubMed:28439027,
FT ECO:0007744|PDB:5B4E"
FT BINDING 161
FT /ligand="ATP"
FT /ligand_id="ChEBI:CHEBI:30616"
FT /evidence="ECO:0000269|PubMed:28439027,
FT ECO:0007744|PDB:5B4E"
FT BINDING 222
FT /ligand="[4Fe-4S] cluster"
FT /ligand_id="ChEBI:CHEBI:49883"
FT /evidence="ECO:0000269|PubMed:28439027,
FT ECO:0007744|PDB:5B4E, ECO:0007744|PDB:5B4F"
FT BINDING 274
FT /ligand="Zn(2+)"
FT /ligand_id="ChEBI:CHEBI:29105"
FT /ligand_label="2"
FT /evidence="ECO:0000269|PubMed:28439027,
FT ECO:0007744|PDB:5B4E, ECO:0007744|PDB:5B4F,
FT ECO:0007744|PDB:5GHA"
FT BINDING 277
FT /ligand="Zn(2+)"
FT /ligand_id="ChEBI:CHEBI:29105"
FT /ligand_label="2"
FT /evidence="ECO:0000269|PubMed:28439027,
FT ECO:0007744|PDB:5B4E, ECO:0007744|PDB:5B4F,
FT ECO:0007744|PDB:5GHA"
FT BINDING 286
FT /ligand="Zn(2+)"
FT /ligand_id="ChEBI:CHEBI:29105"
FT /ligand_label="2"
FT /evidence="ECO:0000269|PubMed:28439027,
FT ECO:0007744|PDB:5B4E, ECO:0007744|PDB:5B4F,
FT ECO:0007744|PDB:5GHA"
FT BINDING 289
FT /ligand="Zn(2+)"
FT /ligand_id="ChEBI:CHEBI:29105"
FT /ligand_label="2"
FT /evidence="ECO:0000269|PubMed:28439027,
FT ECO:0007744|PDB:5B4E, ECO:0007744|PDB:5B4F,
FT ECO:0007744|PDB:5GHA"
FT CROSSLNK 137
FT /note="Glycyl lysine isopeptide (Lys-Gly) (interchain with
FT G-Cter in TtuB)"
FT /evidence="ECO:0000269|PubMed:22467871"
FT CROSSLNK 226
FT /note="Glycyl lysine isopeptide (Lys-Gly) (interchain with
FT G-Cter in TtuB)"
FT /evidence="ECO:0000269|PubMed:22467871"
FT CROSSLNK 229
FT /note="Glycyl lysine isopeptide (Lys-Gly) (interchain with
FT G-Cter in TtuB)"
FT /evidence="ECO:0000269|PubMed:22467871"
FT MUTAGEN 58
FT /note="K->A: About 40% decrease in catalytic activity."
FT /evidence="ECO:0000269|PubMed:23444054"
FT MUTAGEN 59
FT /note="D->A: Complete loss of catalytic activity."
FT /evidence="ECO:0000269|PubMed:23444054"
FT MUTAGEN 130
FT /note="C->S: Decrease in Fe content, and about 90% decrease
FT in catalytic activity."
FT /evidence="ECO:0000269|PubMed:23444054,
FT ECO:0000269|PubMed:28439027"
FT MUTAGEN 133
FT /note="C->S: Decrease in Fe content, and almost complete
FT loss of catalytic activity."
FT /evidence="ECO:0000269|PubMed:23444054,
FT ECO:0000269|PubMed:28439027"
FT MUTAGEN 157
FT /note="H->A: Slight decrease in catalytic activity."
FT /evidence="ECO:0000269|PubMed:23444054"
FT MUTAGEN 158
FT /note="N->A: Slight decrease in catalytic activity."
FT /evidence="ECO:0000269|PubMed:23444054"
FT MUTAGEN 161
FT /note="D->A: Complete loss of catalytic activity."
FT /evidence="ECO:0000269|PubMed:23444054"
FT MUTAGEN 203
FT /note="E->A: About 80% decrease in catalytic activity."
FT /evidence="ECO:0000269|PubMed:23444054"
FT MUTAGEN 222
FT /note="C->S: Decrease in Fe content, and almost complete
FT loss of catalytic activity."
FT /evidence="ECO:0000269|PubMed:23444054,
FT ECO:0000269|PubMed:28439027"
FT TURN 4..6
FT /evidence="ECO:0007829|PDB:5GHA"
FT STRAND 11..14
FT /evidence="ECO:0007829|PDB:5GHA"
FT TURN 15..18
FT /evidence="ECO:0007829|PDB:5GHA"
FT STRAND 19..21
FT /evidence="ECO:0007829|PDB:5GHA"
FT HELIX 23..40
FT /evidence="ECO:0007829|PDB:5GHA"
FT STRAND 49..53
FT /evidence="ECO:0007829|PDB:5GHA"
FT HELIX 58..69
FT /evidence="ECO:0007829|PDB:5GHA"
FT STRAND 73..80
FT /evidence="ECO:0007829|PDB:5GHA"
FT HELIX 84..99
FT /evidence="ECO:0007829|PDB:5GHA"
FT TURN 100..102
FT /evidence="ECO:0007829|PDB:5GHA"
FT STRAND 105..109
FT /evidence="ECO:0007829|PDB:5GHA"
FT HELIX 110..114
FT /evidence="ECO:0007829|PDB:5GHA"
FT HELIX 118..125
FT /evidence="ECO:0007829|PDB:5GHA"
FT HELIX 129..147
FT /evidence="ECO:0007829|PDB:5GHA"
FT STRAND 151..154
FT /evidence="ECO:0007829|PDB:5GHA"
FT HELIX 159..171
FT /evidence="ECO:0007829|PDB:5GHA"
FT STRAND 183..185
FT /evidence="ECO:0007829|PDB:5B4E"
FT STRAND 193..195
FT /evidence="ECO:0007829|PDB:5GHA"
FT TURN 197..200
FT /evidence="ECO:0007829|PDB:5GHA"
FT HELIX 203..213
FT /evidence="ECO:0007829|PDB:5GHA"
FT STRAND 225..227
FT /evidence="ECO:0007829|PDB:5B4E"
FT HELIX 229..243
FT /evidence="ECO:0007829|PDB:5GHA"
FT HELIX 247..258
FT /evidence="ECO:0007829|PDB:5GHA"
FT HELIX 260..262
FT /evidence="ECO:0007829|PDB:5GHA"
FT TURN 275..277
FT /evidence="ECO:0007829|PDB:5GHA"
FT STRAND 280..285
FT /evidence="ECO:0007829|PDB:5GHA"
FT HELIX 287..301
FT /evidence="ECO:0007829|PDB:5GHA"
SQ SEQUENCE 321 AA; 36194 MW; 2DB12E870BFA5314 CRC64;
MVCKVCGQKA QVEMRSRGLA LCREHYLDWF VKETERAIRR HRMLLPGERV LVAVSGGKDS
LALWDVLSRL GYQAVGLHIE LGIGEYSKRS LEVTQAFARE RGLELLVVDL KEAYGFGVPE
LARLSGRVAC SACGLSKRYI INQVAVEEGF RVVATGHNLD DEAAVLFGNL LNPQEETLSR
QGPVLPEKPG LAARVKPFYR FSEREVLSYT LLRGIRYLHE ECPNAKGAKS LLYKEALNLV
ERSMPGAKLR FLDGFLEKIR PRLDVGEEVA LRECERCGYP TTGAVCAFCR MWDAVYRRAK
KRKLLPEEVS FRPRVKPLRA G
