Literature summary for 2.1.1.200 extracted from

Jaroensuk, J.; Atichartpongkul, S.; Chionh, Y.; Hwa Wong, Y.; Liew, C.; McBee, M.; Thongdee, N.; Prestwich, E.; DeMott, M.; Mongkolsuk, S.; Dedon, P.; Lescar, J.; Fuangthong, M.
Methylation at position 32 of tRNA catalyzed by TrmJ alters oxidative stress response in Pseudomonas aeruginosa (2016), Nucleic Acids Res., 44, 10834-10848 .

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Cloned(Commentary)

Cloned (Comment)	Organism
gene trmJ, recombinant expression of His-tagged complete TrmJ protein spanning residues 1-257 (TrmJ) and of His-tagged N-terminal SAM-binding domain, residues 1-167, in Escherichia coli strain BL21(DE3) Rosetta T1R, complementation of a trmJ mutant strain by expression of the endogenous full-length enzyme	Pseudomonas aeruginosa

Crystallization (Commentary)

Crystallization (Comment)	Organism
recombinant TrmJ-NTD protein, hanging drop vapour diffusion method, mixing of 0.001 ml of 20 mg/ml protein solution with 0.002 ml of reservoir solution containing Bis-Tris propane, pH 6.5, 0.2 M ammonium chloride, 25% w/v PEG 3350, 20°C, or with 0.03 M MgCl2, 0.03 M CaCl2, 20% w/v PEG MME550, 10% w/v PEG 20000, and 0.1 M MOPS/Na-HEPES, pH 7.5, and soaking with 1 mM sinefungin, 20°C, resulting in two different crystal forms, method optimization, X-ray diffraction structure determination and analysis at 1.7 A and 1.8-2.2 A resolution, respectively	Pseudomonas aeruginosa

Crystallization (Comment)

Organism

recombinant TrmJ-NTD protein, hanging drop vapour diffusion method, mixing of 0.001 ml of 20 mg/ml protein solution with 0.002 ml of reservoir solution containing Bis-Tris propane, pH 6.5, 0.2 M ammonium chloride, 25% w/v PEG 3350, 20°C, or with 0.03 M MgCl2, 0.03 M CaCl2, 20% w/v PEG MME550, 10% w/v PEG 20000, and 0.1 M MOPS/Na-HEPES, pH 7.5, and soaking with 1 mM sinefungin, 20°C, resulting in two different crystal forms, method optimization, X-ray diffraction structure determination and analysis at 1.7 A and 1.8-2.2 A resolution, respectively

Pseudomonas aeruginosa

Inhibitors

Inhibitors	Comment	Organism	Structure
sinefungin	SAM analogue, enzyme binding structure analysis from complex crystal structure, structure of TrmJ-NTD bound to sinefungin, overview	Pseudomonas aeruginosa

Metals/Ions

Metals/Ions	Comment	Organism	Structure
Mg2+	required	Pseudomonas aeruginosa

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
S-adenosyl-L-methionine + cytidine32 in tRNA	Pseudomonas aeruginosa	-	S-adenosyl-L-homocysteine + 2'-O-methylcytidine32 in tRNA	-	?
S-adenosyl-L-methionine + cytidine32 in tRNA	Pseudomonas aeruginosa UCBPP-PA14	-	S-adenosyl-L-homocysteine + 2'-O-methylcytidine32 in tRNA	-	?

Organism

Organism	UniProt	Comment	Textmining
Pseudomonas aeruginosa	A0A0H2ZF87	-	-
Pseudomonas aeruginosa UCBPP-PA14	A0A0H2ZF87	-	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant His-tagged complete TrmJ protein spanning residues 1-257 (TrmJ) and His-tagged N-terminal SAM-binding domain, residues 1-167, from Escherichia coli strain BL21(DE3) Rosetta T1R by nickel affinity chromatography and two different steps of gel filtration	Pseudomonas aeruginosa

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
additional information	PA14 TrmJ catalyzes 2'-O-methylation of C, U and A at position 32 in the tRNA anticodon loop. tRNA substrates for reaction with PA14 TrmJ are prepared by in vitro T7 transcription. Substrate specificity, overview. No or poor activity with tRNASer(UGA) and tRNAHis(GUG). Structure analysis of TrmJ-NTD tRNA binding	Pseudomonas aeruginosa	?	-	-
additional information	PA14 TrmJ catalyzes 2'-O-methylation of C, U and A at position 32 in the tRNA anticodon loop. tRNA substrates for reaction with PA14 TrmJ are prepared by in vitro T7 transcription. Substrate specificity, overview. No or poor activity with tRNASer(UGA) and tRNAHis(GUG). Structure analysis of TrmJ-NTD tRNA binding	Pseudomonas aeruginosa UCBPP-PA14	?	-	-
S-adenosyl-L-methionine + adenine32 in tRNAPro(GGG)	tRNAPro(GGG) is the most efficient substrate, with methyladenine formation an order of magnitude higher than methyluridine or methylcytidine in other substrates	Pseudomonas aeruginosa	S-adenosyl-L-homocysteine + 2'-O-methyladenine32 in tRNAPro(GGG)	-	?
S-adenosyl-L-methionine + adenine32 in tRNAPro(GGG)	tRNAPro(GGG) is the most efficient substrate, with methyladenine formation an order of magnitude higher than methyluridine or methylcytidine in other substrates	Pseudomonas aeruginosa UCBPP-PA14	S-adenosyl-L-homocysteine + 2'-O-methyladenine32 in tRNAPro(GGG)	-	?
S-adenosyl-L-methionine + cytidine32 in tRNA	-	Pseudomonas aeruginosa	S-adenosyl-L-homocysteine + 2'-O-methylcytidine32 in tRNA	-	?
S-adenosyl-L-methionine + cytidine32 in tRNA	-	Pseudomonas aeruginosa UCBPP-PA14	S-adenosyl-L-homocysteine + 2'-O-methylcytidine32 in tRNA	-	?
S-adenosyl-L-methionine + cytidine32 in tRNAMet(CAU)	-	Pseudomonas aeruginosa	S-adenosyl-L-homocysteine + 2'-O-methylcytisine32 in tRNA	-	?
S-adenosyl-L-methionine + cytidine32 in tRNAMet(CAU)	-	Pseudomonas aeruginosa UCBPP-PA14	S-adenosyl-L-homocysteine + 2'-O-methylcytisine32 in tRNA	-	?
S-adenosyl-L-methionine + cytidine32 in tRNATrp(CCA)	-	Pseudomonas aeruginosa	S-adenosyl-L-homocysteine + 2'-O-methylcytidine32 in tRNATrp(CCA)	-	?
S-adenosyl-L-methionine + cytidine32 in tRNATrp(CCA)	-	Pseudomonas aeruginosa UCBPP-PA14	S-adenosyl-L-homocysteine + 2'-O-methylcytidine32 in tRNATrp(CCA)	-	?
S-adenosyl-L-methionine + uridine32 in tRNAGln(UUG)	-	Pseudomonas aeruginosa	S-adenosyl-L-homocysteine + 2'-O-methyluridine32 in tRNAGln(UUG)	-	?
S-adenosyl-L-methionine + uridine32 in tRNAPro(CGG)	-	Pseudomonas aeruginosa	S-adenosyl-L-homocysteine + 2'-O-methyluridine32 in tRNAPro(CGG)	-	?
S-adenosyl-L-methionine + uridine32 in tRNAPro(UGG)	-	Pseudomonas aeruginosa	S-adenosyl-L-homocysteine + 2'-O-methyluridine32 in tRNAPro(UGG)	-	?

Subunits

Subunits	Comment	Organism
dimer	-	Pseudomonas aeruginosa
More	the free catalytic N-terminal domain of TrmJ shows a 2fold symmetrical dimer with an active site located at the interface between the monomers and a flexible basic loop positioned to bind tRNA, with conformational changes upon binding of the SAM-analogue sinefungin. Analysis of the structure of the N-terminal domain of TrmJ, overview. Residues 166-GKPTKMEK-173 connect the N- to the C-terminal domain of TrmJ in a flexible way in the absence of a tRNA ligand	Pseudomonas aeruginosa

Synonyms

Synonyms	Comment	Organism
2'-O-tRNA methyltransferase	-	Pseudomonas aeruginosa
PA14 14690	-	Pseudomonas aeruginosa
TrmJ	-	Pseudomonas aeruginosa
tRNA (cytidine(32)/uridine(32)/adenosine(32)-2'-O)-methyltransferase	-	Pseudomonas aeruginosa
tRNA:Cm32/Um32/Am32 methyltransferase	-	Pseudomonas aeruginosa

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
37	-	assay at	Pseudomonas aeruginosa

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
7.5	-	assay at	Pseudomonas aeruginosa

Cofactor

Cofactor	Comment	Organism	Structure
S-adenosyl-L-methionine	SAM	Pseudomonas aeruginosa

General Information

General Information	Comment	Organism
evolution	PA14 14690 possesses the conserved region of TrmJ-specific motif, a consensus TXARXR sequence (residues 79-84). This motif has been shown to be critical for binding of SAM/SAH and tRNA. PA14 14690 also contains a conserved Arg at position 23, which is proposed to be involved in its catalytic activity. Another catalytic residue, the Tyr at position 141, is replaced by Phe. In addition to Arg positions 82 and 84 in the TrmJ-specific motif, PA14 14690 also contains more key conserved tRNA binding residues including arginine at positions 100, 101 and 105	Pseudomonas aeruginosa
malfunction	loss of trmJ causes an increased susceptibility to H2O2. Phenotypic analysis of cytotoxicity induced by H2O2 in PA14 wild-type strain, trmJ mutant strain, and the trmJ complementation strain or trmJC, overview. Inactivation of trmJ abolishes Cm, Um, or Am formation in Pseudomonas aeruginosa tRNA	Pseudomonas aeruginosa
metabolism	methylation at position 32 of tRNA catalyzed by TrmJ alters oxidative stress response in Pseudomonas aeruginosa	Pseudomonas aeruginosa
additional information	semi-quantitative real-time PCR analysis of oxyR, katA, katB, katE, ankB and recG transcripts in response to H2O2. Analysis of ribonucleosides by HPLC-coupled tandem quadrupole mass spectrometry	Pseudomonas aeruginosa
physiological function	PA14 trmJ encodes a tRNA (cytidine(32)/uridine(32)/adenosine(32)-2'-O)-methyltransferase that regulates the expression of oxidative stress response genes. Role of tRNA modifications in the oxidative stress response of prokaryotes. TrmJ is a tRNA:Cm32/Um32/Am32 methyltransferase involved in translational fidelity and the oxidative stress response. TrmJ confers H2O2 resistance in part by upregulating the expression of catalase genes	Pseudomonas aeruginosa