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Literature summary extracted from

  • Shao, Z.; Yan, W.; Peng, J.; Zuo, X.; Zou, Y.; Li, F.; Gong, D.; Ma, R.; Wu, J.; Shi, Y.; Zhang, Z.; Teng, M.; Li, X.; Gong, Q.
    Crystal structure of tRNA m1G9 methyltransferase Trm10: insight into the catalytic mechanism and recognition of tRNA substrate (2014), Nucleic Acids Res., 42, 509-525.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

EC Number Cloned (Comment) Organism
2.1.1.221 recombinant expression of N-terminally His-tagged enzyme Trm10 in Escherichia coli strain BL21/Gold(DE3) Saccharomyces cerevisiae
2.1.1.221 recombinant expression of N-terminally His-tagged enzyme Trm10 in Escherichia coli strain BL21/Gold(DE3), selenomethionine-labeled spTrm10_74 (residues 74-281) is expressed in Escherichia coli strain B834, recombinant wild-type and mutant Q226A TrmT10C (residues 40-403) is coexpressed with His-tagged protein SDR5C1 Schizosaccharomyces pombe

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
2.1.1.221 purified enzyme free or in complex with S-adenosyl-L-homocysteine, X-ray diffraction structure determination and analysis at 2.0-2.5 A resolution, molecular replacement Schizosaccharomyces pombe
2.1.1.221 purified enzyme in complex with S-adenosyl-L-homocysteine, X-ray diffraction structure determination and analysis at 1.8 A resolution, molecular replacement Saccharomyces cerevisiae

Protein Variants

EC Number Protein Variants Comment Organism
2.1.1.221 D207N site-directed mutagenesis, inactive mutant Schizosaccharomyces pombe
2.1.1.221 D210N site-directed mutagenesis, inactive mutant Saccharomyces cerevisiae
2.1.1.221 K110E/R121E/R127E site-directed mutagenesis, inactive mutant Schizosaccharomyces pombe
2.1.1.221 K153E/R147E site-directed mutagenesis, inactive mutant Schizosaccharomyces pombe
2.1.1.221 K208A site-directed mutagenesis, mutation of a nucleoside binding pocket residue, the mutant shows 28% reduced activity compared to the wild-type enzyme Schizosaccharomyces pombe
2.1.1.221 N209A site-directed mutagenesis, the mutant shows increased Km for tRNA compared to the wild-type Schizosaccharomyces pombe
2.1.1.221 N212A site-directed mutagenesis, the mutant shows increased Km for tRNA compared to the wild-type Saccharomyces cerevisiae
2.1.1.221 Q118A site-directed mutagenesis, mutation of a nucleoside binding pocket residue, inactive mutant Schizosaccharomyces pombe
2.1.1.221 T244A site-directed mutagenesis, mutation of a nucleoside binding pocket residue, the mutant shows 65% reduced activity compared to the wild-type enzyme Schizosaccharomyces pombe
2.1.1.221 V206A site-directed mutagenesis, mutation of a nucleoside binding pocket residue, the mutant shows 81% reduced activity compared to the wild-type enzyme Schizosaccharomyces pombe

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
2.1.1.221 additional information
-
 additional information kinetic mechanism of Trm10, overview Schizosaccharomyces pombe
2.1.1.221 additional information
-
 additional information kinetic mechanism of Trm10, overview Saccharomyces cerevisiae
2.1.1.221 0.0032
-
 guanine9 in tRNAGly tRNA from Schizosaccharomyces pombe, recombinant enzyme, pH and temperature not specified in the publication Schizosaccharomyces pombe
2.1.1.221 0.0053
-
 guanine9 in tRNAGly tRNA from Schizosaccharomyces pombe, recombinant enzyme, pH and temperature not specified in the publication Saccharomyces cerevisiae

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
2.1.1.221 S-adenosyl-L-methionine + guanine9 in tRNAGly Schizosaccharomyces pombe Trm10 is specific for G9 of tRNAGly S-adenosyl-L-homocysteine + N1-methylguanine9 in tRNAGly
-
 ?
2.1.1.221 S-adenosyl-L-methionine + guanine9 in tRNAGly Saccharomyces cerevisiae Trm10 is specific for G9 of tRNAGly S-adenosyl-L-homocysteine + N1-methylguanine9 in tRNAGly
-
 ?
2.1.1.221 S-adenosyl-L-methionine + guanine9 in tRNAGly Schizosaccharomyces pombe 972 Trm10 is specific for G9 of tRNAGly S-adenosyl-L-homocysteine + N1-methylguanine9 in tRNAGly
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
2.1.1.221 Saccharomyces cerevisiae Q12400
-
-
2.1.1.221 Schizosaccharomyces pombe O14214
-
-
2.1.1.221 Schizosaccharomyces pombe 972 O14214
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
2.1.1.221 recombinant N-terminally His-tagged enzyme Trm10 from Escherichia coli strain BL21/Gold(DE3) by nickel affinity chromatography, gel filtration, and anion exchange chromatography, followed by dialysis Saccharomyces cerevisiae
2.1.1.221 recombinant N-terminally His-tagged enzyme Trm10 from Escherichia coli strain BL21/Gold(DE3) by nickel affinity chromatography, gel filtration, and anion exchange chromatography, followed by dialysis. Purification of the recombinant TrmT10C-SDR5C1 wild-type and mutant complexes by nickel affinity chromatography Schizosaccharomyces pombe

Reaction

EC Number Reaction Comment Organism Reaction ID
2.1.1.221 S-adenosyl-L-methionine + guanine9 in tRNA = S-adenosyl-L-homocysteine + N1-methylguanine9 in tRNA catalytic mechanism and recognition of tRNA substrate, overview. The N-terminal extension of Trm10 is necessary for substrate tRNA recognition Schizosaccharomyces pombe
a
2.1.1.221 S-adenosyl-L-methionine + guanine9 in tRNA = S-adenosyl-L-homocysteine + N1-methylguanine9 in tRNA catalytic mechanism and recognition of tRNA substrate, overview. The N-terminal extension of Trm10 is necessary for substrate tRNA recognition Saccharomyces cerevisiae
a

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2.1.1.221 additional information no activity of the enzyme with three sptRNAGly mutants at Position 9 (G9A, G9C, and G9U) Schizosaccharomyces pombe ?
-
 ?
2.1.1.221 additional information no activity of the enzyme with three sptRNAGly mutants at Position 9 (G9A, G9C, and G9U) Saccharomyces cerevisiae ?
-
 ?
2.1.1.221 additional information no activity of the enzyme with three sptRNAGly mutants at Position 9 (G9A, G9C, and G9U) Schizosaccharomyces pombe 972 ?
-
 ?
2.1.1.221 S-adenosyl-L-methionine + guanine9 in tRNAGly Trm10 is specific for G9 of tRNAGly Schizosaccharomyces pombe S-adenosyl-L-homocysteine + N1-methylguanine9 in tRNAGly
-
 ?
2.1.1.221 S-adenosyl-L-methionine + guanine9 in tRNAGly Trm10 is specific for G9 of tRNAGly Saccharomyces cerevisiae S-adenosyl-L-homocysteine + N1-methylguanine9 in tRNAGly
-
 ?
2.1.1.221 S-adenosyl-L-methionine + guanine9 in tRNAGly Trm10 is specific for G9 of tRNAGly Schizosaccharomyces pombe 972 S-adenosyl-L-homocysteine + N1-methylguanine9 in tRNAGly
-
 ?

Subunits

EC Number Subunits Comment Organism
2.1.1.221 monomer the enzyme adopts a globular alpha/beta structure consisting of six-stranded beta-sheets sandwiched by alpha-helices at both sides, small angle X-ray scattering analysis Schizosaccharomyces pombe
2.1.1.221 monomer the enzyme adopts a globular alpha/beta structure consisting of six-stranded beta-sheets sandwiched by alpha-helices at both sides, small angle X-ray scattering analysis Saccharomyces cerevisiae

Synonyms

EC Number Synonyms Comment Organism
2.1.1.221 Trm10
-
 Schizosaccharomyces pombe
2.1.1.221 Trm10
-
 Saccharomyces cerevisiae
2.1.1.221 tRNA m1G9 methyltransferase
-
 Schizosaccharomyces pombe
2.1.1.221 tRNA m1G9 methyltransferase
-
 Saccharomyces cerevisiae
2.1.1.221 tRNA m1G9 MTase
-
 Schizosaccharomyces pombe
2.1.1.221 tRNA m1G9 MTase
-
 Saccharomyces cerevisiae

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
2.1.1.221 0.0027
-
 guanine9 in tRNAGly tRNA from Schizosaccharomyces pombe, recombinant enzyme, pH and temperature not specified in the publication Schizosaccharomyces pombe
2.1.1.221 0.0028
-
 guanine9 in tRNAGly tRNA from Schizosaccharomyces pombe, recombinant enzyme, pH and temperature not specified in the publication Saccharomyces cerevisiae

Cofactor

EC Number Cofactor Comment Organism Structure
2.1.1.221 S-adenosyl-L-methionine binding structure, overview Schizosaccharomyces pombe
2.1.1.221 S-adenosyl-L-methionine binding structure, overview Saccharomyces cerevisiae
2.1.1.221 SDR5C1 protein enzyme TrmT10C requires another protein, SDR5C1 (MRPP2), to execute its m1R9 (G9 and A9) MTase and additional RNase P activity Schizosaccharomyces pombe

General Information

EC Number General Information Comment Organism
2.1.1.221 evolution the enzyme Trm10 belongs to the SPOUT superfamily. Trm10 behaves as a monomer in solution, whereas other members of the SPOUT superfamily all function as homodimers. The MTase domain (the catalytic domain) of the Trm10 family displays a typical SpoU-TrmD (SPOUT) fold. Trm10 from Schizosaccharomyces pombe demonstrates identical tRNA MTase activity as Trm10 from Saccharomyces cerevisiae Schizosaccharomyces pombe
2.1.1.221 evolution the enzyme Trm10 belongs to the SPOUT superfamily. Trm10 behaves as a monomer in solution, whereas other members of the SPOUT superfamily all function as homodimers. The MTase domain (the catalytic domain) of the Trm10 family displays a typical SpoU-TrmD (SPOUT) fold. Trm10 from Schizosaccharomyces pombe demonstrates identical tRNA MTase activity as Trm10 from Saccharomyces cerevisiae Saccharomyces cerevisiae
2.1.1.221 additional information enzyme overall structure analysis, active site structure, overview Schizosaccharomyces pombe
2.1.1.221 additional information enzyme overall structure analysis, active site structure, overview Saccharomyces cerevisiae

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
2.1.1.221 0.538
-
 guanine9 in tRNAGly tRNA from Schizosaccharomyces pombe, recombinant enzyme, pH and temperature not specified in the publication Saccharomyces cerevisiae
2.1.1.221 1.66
-
 guanine9 in tRNAGly tRNA from Schizosaccharomyces pombe, recombinant enzyme, pH and temperature not specified in the publication Schizosaccharomyces pombe