Information on EC 2.1.1.207 - tRNA (cytidine34-2'-O)-methyltransferase

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The expected taxonomic range for this enzyme is: Escherichia coli

EC NUMBER
COMMENTARY hide
2.1.1.207
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RECOMMENDED NAME
GeneOntology No.
tRNA (cytidine34-2'-O)-methyltransferase
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
S-adenosyl-L-methionine + 5-carboxymethylaminomethyluridine34 in tRNALeu = S-adenosyl-L-homocysteine + 5-carboxymethylaminomethyl-2'-O-methyluridine34 in tRNALeu
show the reaction diagram
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S-adenosyl-L-methionine + cytidine34 in tRNA = S-adenosyl-L-homocysteine + 2'-O-methylcytidine34 in tRNA
show the reaction diagram
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SYSTEMATIC NAME
IUBMB Comments
S-adenosyl-L-methionine:tRNA (cytidine34/5-carboxymethylaminomethyluridine34-2'-O)-methyltransferase
The enzyme from Escherichia coli catalyses the 2'-O-methylation of cytidine or 5-carboxymethylaminomethyluridine at the wobble position at nucleotide 34 in tRNALeuCmAA and tRNALeucmnm5UmAA. The enzyme is selective for the two tRNALeu isoacceptors and only methylates these when they present the correct anticodon loop sequence and modification pattern. Specifically, YibK requires a pyrimidine nucleoside at position 34, it has a clear preference for an adenosine at position 35, and it fails to methylate without prior addition of the N6-(isopentenyl)-2-methylthioadenosine modification at position 37.
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
gene trmL
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Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
malfunction
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inactivation of yibK leads to loss of 2'-O-methylation at position 34 in both tRNALeu(CmAA) and tRNALeu(cmnm5UmAA). Loss of YibK methylation reduces the efficiency of codon–wobble base interaction. Inactivation of yibK has no detectable effect on steady-state growth rate, although a distinct disadvantage is noted in multiple-round, mixed-population growth experiments, suggesting that the ability to recover from the stationary phase is impaired. Methylation is restored in vivo by complementing with a recombinant copy of yibK
metabolism
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modifications at the wobble uridine, U34, of tRNAs reading two degenerate codons ending in purine are complex and result from the activity of two multi-enzyme pathways, the IscSeMnmA and MnmEG pathways, which independently work on positions 2 and 5 of the U34 pyrimidine ring, respectively, and from a third single-step pathway, controlled by TrmL, i.e. YibK, that modifies the 2'-hydroxyl group of the ribose. TrmL occurs as a late step in the maturation of the tRNALeu isoacceptors
physiological function
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TrmL is the prokaryotic methyltransferase that catalyzes the transfer of the methyl group from S-adenosyl-L-methionine to the wobble base of tRNALeuCAA and tRNALeuUAA isoacceptors. This Cm34/Um34 modification affects codon-anticodon interactions and is essential for translational fidelity
additional information
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
S-adenosyl-L-methionine + 5-carboxymethylaminomethyluridine34 in tRNA
S-adenosyl-L-homocysteine + 5-carboxymethylaminomethyl-2'-O-methyluridine34 in tRNA
show the reaction diagram
S-adenosyl-L-methionine + 5-carboxymethylaminomethyluridine34 in tRNALeu
S-adenosyl-L-homocysteine + 5-carboxymethylaminomethyl-2'-O-methyluridine34 in tRNALeu
show the reaction diagram
the enzyme methylates the ribose at the nucleotide 34 wobble position in the two leucyl isoacceptors tRNALeu(CmAA) and tRNALeu(cmnm5UmAA)
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?
S-adenosyl-L-methionine + cytidine34 in tRNA
S-adenosyl-L-homocysteine + 2'-O-methylcytidine34 in tRNA
show the reaction diagram
S-adenosyl-L-methionine + cytidine34 in tRNALeuCAA
S-adenosyl-L-homocysteine + 2'-O-methylcytidine34 in tRNALeuCAA
show the reaction diagram
S-adenosyl-L-methionine + cytidine34 in tRNALeuUAA
S-adenosyl-L-homocysteine + 2'-O-methylcytidine34 in tRNALeuUAA
show the reaction diagram
additional information
?
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NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
S-adenosyl-L-methionine + 5-carboxymethylaminomethyluridine34 in tRNA
S-adenosyl-L-homocysteine + 5-carboxymethylaminomethyl-2'-O-methyluridine34 in tRNA
show the reaction diagram
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TrmL catalyzes the methyl transfer from SAM to the 2'-OH of the wobble nucleotide in tRNALeu cmnm5UmAA, which decodes codons UUA and UUG in the Phe/Leu mixed box, and tRNALeu CmAA, which reads the UUG codon included in the Leu family box
o.e. tRNALeucmnm5UmAA
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?
S-adenosyl-L-methionine + 5-carboxymethylaminomethyluridine34 in tRNALeu
S-adenosyl-L-homocysteine + 5-carboxymethylaminomethyl-2'-O-methyluridine34 in tRNALeu
show the reaction diagram
P0AGJ7
the enzyme methylates the ribose at the nucleotide 34 wobble position in the two leucyl isoacceptors tRNALeu(CmAA) and tRNALeu(cmnm5UmAA)
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?
S-adenosyl-L-methionine + cytidine34 in tRNA
S-adenosyl-L-homocysteine + 2'-O-methylcytidine34 in tRNA
show the reaction diagram
P0AGJ7
the enzyme methylates the ribose at the nucleotide 34 wobble position in the two leucyl isoacceptors tRNALeu(CmAA) and tRNALeu(cmnm5UmAA)
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-
?
S-adenosyl-L-methionine + cytidine34 in tRNALeuCAA
S-adenosyl-L-homocysteine + 2'-O-methylcytidine34 in tRNALeuCAA
show the reaction diagram
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?
S-adenosyl-L-methionine + cytidine34 in tRNALeuUAA
S-adenosyl-L-homocysteine + 2'-O-methylcytidine34 in tRNALeuUAA
show the reaction diagram
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COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
S-adenosyl-L-methionine
additional information
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no requirement of other protein cofactors
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Mg2+
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required
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.00182 - 0.00537
cytidine34 in tRNALeuCAA
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wild-type tRNALeuCAA and diverse mutants thereof, pH 7.5, 37°C
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additional information
additional information
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steady-state kinetic parameters of EcTrmL with modified tRNALeu substrates
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0073 - 0.2
cytidine34 in tRNALeuCAA
Escherichia coli
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wild-type tRNALeuCAA and diverse mutants thereof, pH 7.5, 37°C
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pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7.5
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assay at
8
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assay at
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
PDB
SCOP
CATH
ORGANISM
UNIPROT
Burkholderia pseudomallei (strain 1710b)
Escherichia coli (strain K12)
Escherichia coli (strain K12)
Escherichia coli (strain UTI89 / UPEC)
Haemophilus influenzae (strain ATCC 51907 / DSM 11121 / KW20 / Rd)
Haemophilus influenzae (strain ATCC 51907 / DSM 11121 / KW20 / Rd)
Thermus thermophilus (strain HB27 / ATCC BAA-163 / DSM 7039)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
17700
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x * 17700, TrmL
17726
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2 * 17726, calulated from sequence
19800
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2 * 19800, SDS-PAGE and gel filtration, the overall structure of an EcTrmL monomer subunit is composed of six beta-strands and six alpha-helices, in the order beta1-alpha1-beta2-alpha2-alpha3-beta3-alpha4-beta4-beta5-alpha5-beta6-alpha6. EcTrmL dimer formation is essential for tRNA recognition. The residue Y142 is critical for maintaining the dimeric form of EcTrmL, which is consistent with its central position at the interface
36000
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gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
?
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x * 17700, TrmL
dimer
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2 * 17726, calulated from sequence
homodimer
additional information
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TrmL-catalyzed 2'-O-methylation requires its homodimerization
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
purified enzyme TrmL in apoform and in complex with S-adenosyl-homocysteine, X-ray diffraction structure determination and analysis at 2.0 A resolution, molecular replacement using the structure of Haemophilus influenzae Yibk, PDB ID 1J85 as starting search model, modeling
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity chromatography and gel filtration
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recombinant His-YibK protein
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
gene rmL, recombinant expression of wild-type and mutant enzymes
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gene trmL, recombinant expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3)
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
K42E
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site-directed mutagenesis, inactive mutant
K81E
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site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme
R104E
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site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme
R129E
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site-directed mutagenesis, inactive mutant
R20E
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site-directed mutagenesis, inactive mutant
R28E
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site-directed mutagenesis, the mutant shows slightly increased activity compared to the wild-type enzyme
R43E
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site-directed mutagenesis, inactive mutant
R45E
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site-directed mutagenesis, inactive mutant
R46E
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site-directed mutagenesis, inactive mutant
R59E
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site-directed mutagenesis, inactive mutant
R64E
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site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme
R74E
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site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme
Y142A
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site-directed mutagenesis, the mutant is a monomer in contrast to the wild-type enzyme. Mutant Y142A fails to form a complex with tRNA
K42E
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site-directed mutagenesis, inactive mutant
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K81E
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site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme
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R46E
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site-directed mutagenesis, inactive mutant
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R59E
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site-directed mutagenesis, inactive mutant
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additional information
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