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Literature summary for 2.1.1.228 extracted from

  • Wang, C.; Jia, Q.; Chen, R.; Wei, Y.; Li, J.; Ma, J.; Xie, W.
    Crystal structures of the bifunctional tRNA methyltransferase Trm5a (2016), Sci. Rep., 6, 33553 .
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

Cloned (Comment) Organism
gene trm5a, sequence comparisons, recombinant expression of N-terminally His6-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3) Pyrococcus abyssi

Crystallization (Commentary)

Crystallization (Comment) Organism
purified recombinant PaTrm5a in apo form and in complex with various SAM analogues, mixing PaTrm5a with 1.5 mM SAH or SAM at a protein: ligand molar ratio of 1:3, sitting drop vapor diffusion method, mixing 20 mg/ml protein solution in a 1:1 ratio with well solution containing w/v PEG 3350, 100 mM HEPES, pH 7.5, 100 mM Ca(OAc)2 and 100 mM KCl, method optimmization, 25°C, X-ray diffraction structure determination and analysis at 1.76-2.20 A resolution Pyrococcus abyssi

Protein Variants

Protein Variants Comment Organism
C301S/C308S/C326S site-directed mutagenesis Pyrococcus abyssi
V21C/C301S/C308S/K314C/C326S site-directed mutagenesis Pyrococcus abyssi

Metals/Ions

Metals/Ions Comment Organism Structure
Mg2+ required Pyrococcus abyssi

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
additional information Pyrococcus abyssi bifunctional Trm5a from Pyrococcus abyssi (PaTrm5a) catalyses not only the methylation of N1, but also the further methylation of C7 on 4 demethylwyosine at position 37 to produce isowyosine (EC 2.1.1.228 and EC 2.1.1.282, respectively) ?
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S-adenosyl-L-methionine + guanine37 in tRNAPhe Pyrococcus abyssi
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S-adenosyl-L-homocysteine + N1-methylguanine37 in tRNAPhe
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?

Organism

Organism UniProt Comment Textmining
Pyrococcus abyssi Q9V2G1
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Purification (Commentary)

Purification (Comment) Organism
recombinant N-terminally His6-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity and heparin affinity chromatography, and dialysis Pyrococcus abyssi

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
additional information bifunctional Trm5a from Pyrococcus abyssi (PaTrm5a) catalyses not only the methylation of N1, but also the further methylation of C7 on 4 demethylwyosine at position 37 to produce isowyosine (EC 2.1.1.228 and EC 2.1.1.282, respectively) Pyrococcus abyssi ?
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additional information substrate-binding modes of PaTrm5a, and recognition of substrate analogues, overview Pyrococcus abyssi ?
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S-adenosyl-L-methionine + guanine37 in tRNAPhe
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Pyrococcus abyssi S-adenosyl-L-homocysteine + N1-methylguanine37 in tRNAPhe
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?

Subunits

Subunits Comment Organism
? x * 38500, recombinant N-terminally His6-tagged enzyme, SDS-PAGE Pyrococcus abyssi
More enzyme structure analysis, the 38.5-kDa protein is composed of three domains: D1 (M1-P60), D2 (N70-S162) and D3 (K163-S333) D1 and D2 are connected through an interdomain linker (M61-K69), overview Pyrococcus abyssi

Synonyms

Synonyms Comment Organism
More see also EC 2.1.1.282 Pyrococcus abyssi
PaTrm5a
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Pyrococcus abyssi
TAW22
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Pyrococcus abyssi
Trm5a
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Pyrococcus abyssi

Cofactor

Cofactor Comment Organism Structure
S-adenosyl-L-methionine
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Pyrococcus abyssi

General Information

General Information Comment Organism
evolution during the evolutionary process, some euryarchaeota like Thermococcus and Pyrococcus preserved both the trm5 genes from the crenarchaeal origin as well as the native copy, but others apparently lost the latter. Phylogenetic distribution analyses of trm5 homologues in archaeal genomes allow the identification of three archaeal Trm5 (aTrm5) subfamilies: Trm5a, Trm5b, and Trm5c. Trm5b refers to the native form, while Trm5a refers to the crenarchaeal origin, and Trm5c to other members with divergent Trm5 sequences11. The three Trm5s differ substantially in primary sequences Pyrococcus abyssi
metabolism the enzyme is part of the The biosynthetic pathway of mimG in Pyrococcus abyssi, overview. In archaea, G37 hypermodification in tRNAPhe leads to wyosine derivatives. They are important in reading-frame maintenance during protein synthesis, while the absence of such modifications results in elevated error rates in +1 frame-shifting. Among the modification products, 7-methylwyosine (mimG) is perhaps the earliest and minimalist version of the wyosine derivatives unique to some archaea, and 4-demethylwyosine (imG-14), isowyosine (imG2) have also been identified as intermediates along the pathway. The first biosynthetic step of mimG is the formation of m1G37, catalysed by the S-adenosine-L-methionine (SAM)-dependent tRNA methyltransferase named Trm5, which belongs to class-I methyltransferases. The second step is the complex radical-mediated formation of imG-14, catalyzed by the radical SAM enzyme Taw1. The Trm5 enzyme from the archaeon Pyrococcus abyssi (PaTrm5a) also catalyzes the methylation of C7 on imG-14 to produce imG2 (EC 2.1.1.282), which is further methylated on the N4 position of the imidazo-purine ring by Taw3 to form mimG Pyrococcus abyssi
additional information enzyme structure comparisons Pyrococcus abyssi
physiological function tRNA methyltransferase Trm5 catalyses the transfer of a methyl group from S-adenosyl-L-methionine to G37 in eukaryotes and archaea. The N1-methylated guanosine is the product of the initial step of the wyosine hypermodification, which is essential for the maintenance of the reading frame during translation. As a unique member of this enzyme family, Trm5a from Pyrococcus abyssi (PaTrm5a) catalyses not only the methylation of N1, but also the further methylation of C7 on 4-demethylwyosine at position 37 to produce isowyosine Pyrococcus abyssi