In contrast to the archaeal enzyme tRNA (guanine10-N2)-dimethyltransferase (EC 2.1.1.213), tRNA (guanine10-N2)-methyltransferase from yeast does not catalyse the methylation from N2-methylguanine10 to N2-dimethylguanine10 in tRNA.
The expected taxonomic range for this enzyme is: Eukaryota, Archaea
In contrast to the archaeal enzyme tRNA (guanine10-N2)-dimethyltransferase (EC 2.1.1.213), tRNA (guanine10-N2)-methyltransferase from yeast does not catalyse the methylation from N2-methylguanine10 to N2-dimethylguanine10 in tRNA.
although the eukaryotic counterpart (Trm11) requires another subunit Trm112 for enzymatic activity, the archaeal enzyme does not require a partner subunit
although the eukaryotic counterpart (Trm11) requires another subunit Trm112 for enzymatic activity, the archaeal enzyme does not require a partner subunit
the enzyme is composed of at least two subunits that are associated in vivo: Trm11p, which is the catalytic subunit, and Trm112p, a putative zinc-binding protein
zinc might be important for Trm11 binding to Trm112, most probably by maintaining the three-dimensional structure of Trm112 zinc-binding domain, which in the other Trm112-MTase complexes is directly interacting with the MTase domains
dependent on, Trm11 proves to be completely inactive alone. This lack of activity cannot be attributed to protein unfolding as the circular dichroism (CD) spectrum recorded on this protein is typical of well-folded proteins containing alpha-helices and beta-strands. Trm112 stimulates SAM binding to Trm11 and contributes to tRNA binding. Analysis of the activation mode of the eukaryotic m2G10 tRNA methyltransferase Trm11 by its partner protein Trm112. Yeast Trm112 has a calculated molecular weight of 15067.6 Da. A zinc atom is attached to its Zn-binding domain. The Trm112-Trm11 interaction mode is reminiscent of the other Trm112-MTase complexes. Three-dimensional Trm11-Trm112 complex structrue analysis, and thermodynamics of interaction at 10°C, overview
although the eukaryotic counterpart (Trm11) requires another subunit Trm112 for enzymatic activity, the archaeal enzyme does not require a partner subunit. Furthermore, the eukaryotic Trm11-Trm112 complex catalyzes a single methyl transfer reaction and forms only m2G10 in tRNA. The archaeal enzyme is called Trm-G10 or Trm-m22G10 to distinguish it from the eukaryotic enzyme
although the eukaryotic counterpart (Trm11) requires another subunit Trm112 for enzymatic activity, the archaeal enzyme does not require a partner subunit. Furthermore, the eukaryotic Trm11-Trm112 complex catalyzes a single methyl transfer reaction and forms only m2G10 in tRNA. The archaeal enzyme is called Trm-G10 or Trm-m22G10 to distinguish it from the eukaryotic enzyme
although the eukaryotic counterpart (Trm11) requires another subunit Trm112 for enzymatic activity, the archaeal enzyme does not require a partner subunit. Furthermore, the eukaryotic Trm11-Trm112 complex catalyzes a single methyl transfer reaction and forms only m2G10 in tRNA. The archaeal enzyme is called Trm-G10 or Trm-m22G10 to distinguish it from the eukaryotic enzyme
While deletion of TRM11 has no detectable phenotype under laboratory conditions, deletion of TRM112 leads to a severe growth defect, suggesting that it has additional functions in the cell
TRM112 mutant strains show growth defects, as well as nuclear genomic instability and mitotic defects, overview. Chromosome instability increases 6-7-fold in trm112-0 compared with wild-type. trm112-0 strain is resistant to zymocin, thetRNase toxin produced by Kluyveromyces lactis that specifically targets the mcm5s2U modification at position 34 and cleaves tRNAUUGGln, tRNAUUCGlu, and tRNAUUU
Trm11p and Trm112p are two interacting proteins that are both required for catalyzing the formation of m2G at position 10 in several tRNAs. Trm11p is the catalytic subunit, but also Trm112p is essential for the formation of m2G10, Trm112p is essential for the activity of Trm11p, another tRNA methyltransferase, because Trm112p is capable of directing the proper folding of Trm11p, leading to the synthesis of an active complex. Trm112p is required in vivo for the formation of mcm5U34 and mcm5s2U34, overview. Trm112p is also required for the activity of Mtq2p, a protein methyltransferase that catalyzes the methylation of the glutamine of the universally conserved GGQ tripeptide of the translation termination factor eRF1/Sup45. Trm112p is associated with other partners involved in ribosome biogenesis and chromatin remodeling, suggesting that it has additional roles in the cell
the enzyme is active only in complex with its partner protein Trm112, Trm11 proves to be completely inactive alone. The Trm112-Trm11 interaction mode is reminiscent of the other Trm112-MTase complexes. zinc might be important for Trm11 binding to Trm112, most probably by maintaining the three-dimensional structure of Trm112 zinc-binding domain, which in the other Trm112-MTase complexes is directly interacting with the MTase domains. Trm112 stimulates SAM binding to Trm11 and contributes to tRNA binding. Also the tRNA 3'-CCA tail from the aminoacyl stem loop for tRNA is important for methyltransferase activity
tRNA m2G10/m2 2G10 methyltransferase (archaeal Trm11) methylates the 2-amino group in guanosine at position 10 in tRNA and forms N2,N2-dimethylguanosine (m22G10) via intermediate N2-methylguanosine (m2G10). The archaeal Trm11 is required for Thermococcus kodakarensis survival at high temperatures. The m22G10 modification might have effects on stabilization of tRNA and/or correct folding of tRNA at the high temperatures
tRNA m2G10/m2 2G10 methyltransferase (archaeal Trm11) methylates the 2-amino group in guanosine at position 10 in tRNA and forms N2,N2-dimethylguanosine (m22G10) via intermediate N2-methylguanosine (m2G10). The archaeal Trm11 is required for Thermococcus kodakarensis survival at high temperatures. The m22G10 modification might have effects on stabilization of tRNA and/or correct folding of tRNA at the high temperatures
the enzyme is active only in complex with its partner protein Trm112, Trm11 proves to be completely inactive alone. The Trm112-Trm11 interaction mode is reminiscent of the other Trm112-MTase complexes. zinc might be important for Trm11 binding to Trm112, most probably by maintaining the three-dimensional structure of Trm112 zinc-binding domain, which in the other Trm112-MTase complexes is directly interacting with the MTase domains. Trm112 stimulates SAM binding to Trm11 and contributes to tRNA binding. Also the tRNA 3'-CCA tail from the aminoacyl stem loop for tRNA is important for methyltransferase activity
tRNA m2G10/m2 2G10 methyltransferase (archaeal Trm11) methylates the 2-amino group in guanosine at position 10 in tRNA and forms N2,N2-dimethylguanosine (m22G10) via intermediate N2-methylguanosine (m2G10). The archaeal Trm11 is required for Thermococcus kodakarensis survival at high temperatures. The m22G10 modification might have effects on stabilization of tRNA and/or correct folding of tRNA at the high temperatures
composed of at least two subunits that are associated in vivo: Trm11p (Yol124c), which is the catalytic subunit, and Trm112p (Ynr046w), a putative zinc-binding protein. While deletion of TRM11 has no detectable phenotype under laboratory conditions, deletion of TRM112 leads to a severe growth defect, suggesting that it has additional functions in the cell. Trm112p is associated with at least four proteins: two tRNA methyltransferases (Trm9p and Trm11p), one putative protein methyltransferase (Mtc6p/Ydr140w), and one protein with a Rossmann fold dehydrogenase domain (Lys9p/Ynr050c)
composed of at least two subunits that are associated in vivo: Trm11p (Yol124c), which is the catalytic subunit, and Trm112p (Ynr046w), a putative zinc-binding protein. While deletion of TRM11 has no detectable phenotype under laboratory conditions, deletion of TRM112 leads to a severe growth defect, suggesting that it has additional functions in the cell. Trm112p is associated with at least four proteins: two tRNA methyltransferases (Trm9p and Trm11p), one putative protein methyltransferase (Mtc6p/Ydr140w), and one protein with a Rossmann fold dehydrogenase domain (Lys9p/Ynr050c)
Trm112p is a small 15-kDa protein that contains a zinc finger domain, a motif composed of four cysteines arranged with a certain spacing, which can form a secondary structure resembling a finger
a plasmid containing a wild-type copy of the TRM112 gene (pBL652) is able to restore the formation of the two modified nucleosides in a trm112-0 strain
construction of a gene disruptant trm11 (DELTAtrm11) mutant showing the absence of m2s2G10. The lack of 2-methylguanosine (m2G) at position 67 in the trm11 trm14 double disruptant strain suggests that this methylation is mediated by Trm14, which is an m2G6 methyltransferase. The trm11 gene is reinserted into the chiA (Tk1765, chitinase gene) region in the genomic DNA of the DELTAtrm11 strain. Deletion of the Tk1765 gene does not cause growth defects unless chitin is used as a carbon source. Although its expression level is lower in the complemented strain than in the wild-type strain, Trm11 is expressed in the complemented strain. At 85°C, the wild-type, DELTAtrm11, and complemented strains show similar growth curves. As the temperature increases, the growth of the DELTAtrm11 strain is clearly slower than that of the wild-type or complemented strain. At 95°C, the DELTAtrm11 strain shows a considerable growth defect, whereas the complemented strain grows at approximately the same speed as the wild-type strain, indicating that the growth defect of the DELTAtrm11 strain is due to the lack of archaeal Trm11 protein
construction of a gene disruptant trm11 (DELTAtrm11) mutant showing the absence of m2s2G10. The lack of 2-methylguanosine (m2G) at position 67 in the trm11 trm14 double disruptant strain suggests that this methylation is mediated by Trm14, which is an m2G6 methyltransferase. The trm11 gene is reinserted into the chiA (Tk1765, chitinase gene) region in the genomic DNA of the DELTAtrm11 strain. Deletion of the Tk1765 gene does not cause growth defects unless chitin is used as a carbon source. Although its expression level is lower in the complemented strain than in the wild-type strain, Trm11 is expressed in the complemented strain. At 85°C, the wild-type, DELTAtrm11, and complemented strains show similar growth curves. As the temperature increases, the growth of the DELTAtrm11 strain is clearly slower than that of the wild-type or complemented strain. At 95°C, the DELTAtrm11 strain shows a considerable growth defect, whereas the complemented strain grows at approximately the same speed as the wild-type strain, indicating that the growth defect of the DELTAtrm11 strain is due to the lack of archaeal Trm11 protein
construction of a gene disruptant trm11 (DELTAtrm11) mutant showing the absence of m2s2G10. The lack of 2-methylguanosine (m2G) at position 67 in the trm11 trm14 double disruptant strain suggests that this methylation is mediated by Trm14, which is an m2G6 methyltransferase. The trm11 gene is reinserted into the chiA (Tk1765, chitinase gene) region in the genomic DNA of the DELTAtrm11 strain. Deletion of the Tk1765 gene does not cause growth defects unless chitin is used as a carbon source. Although its expression level is lower in the complemented strain than in the wild-type strain, Trm11 is expressed in the complemented strain. At 85°C, the wild-type, DELTAtrm11, and complemented strains show similar growth curves. As the temperature increases, the growth of the DELTAtrm11 strain is clearly slower than that of the wild-type or complemented strain. At 95°C, the DELTAtrm11 strain shows a considerable growth defect, whereas the complemented strain grows at approximately the same speed as the wild-type strain, indicating that the growth defect of the DELTAtrm11 strain is due to the lack of archaeal Trm11 protein
construction of a gene disruptant trm11 (DELTAtrm11) mutant showing the absence of m2s2G10. The lack of 2-methylguanosine (m2G) at position 67 in the trm11 trm14 double disruptant strain suggests that this methylation is mediated by Trm14, which is an m2G6 methyltransferase. The trm11 gene is reinserted into the chiA (Tk1765, chitinase gene) region in the genomic DNA of the DELTAtrm11 strain. Deletion of the Tk1765 gene does not cause growth defects unless chitin is used as a carbon source. Although its expression level is lower in the complemented strain than in the wild-type strain, Trm11 is expressed in the complemented strain. At 85°C, the wild-type, DELTAtrm11, and complemented strains show similar growth curves. As the temperature increases, the growth of the DELTAtrm11 strain is clearly slower than that of the wild-type or complemented strain. At 95°C, the DELTAtrm11 strain shows a considerable growth defect, whereas the complemented strain grows at approximately the same speed as the wild-type strain, indicating that the growth defect of the DELTAtrm11 strain is due to the lack of archaeal Trm11 protein
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PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
recombinant His-tagged Trm112p, alone or with Trm9p, from Escherichia coli strain S15 (DE3-pLysS) by nickel affinity chromatography and desalting gel filtration
recombinant soluble C-terminally His6-tagged wild-type and mutant enzymes to homogeneity from Escherichia coli strain BL21-Gold (DE3) by nickel affinity chromatography and gel filtration
gene TRM112, phylogenetic analysis, expression of His-tagged Trm112p, alone or with Trm9p, in Escherichia coli strain S15 (DE3-pLysS), Trm112p forms a complex with Trm9p, which renders the latter soluble
gene YOL124c, recombinant expression of soluble C-terminally His6-tagged wild-type and mutant enzymes in Escherichia coli strain BL21-Gold (DE3), coexpression of yeast Trm11 with yeast Trm112 in Escherichia coli, the holonezyme complex is active
recombinant Trm11p expressed in Escherichia coli exhibited no m2G10 formation activity. Trm112p and Trm11p are both required for the formation of m2G10 in vivo as well as in vitro
Hirata, A.; Suzuki, T.; Nagano, T.; Fujii, D.; Okamoto, M.; Sora, M.; Lowe, T.M.; Kanai, T.; Atomi, H.; Suzuki, T.; Hori, H.
Distinct modified nucleosides in tRNATrp from thehyperthermophilic archaeon Thermococcus kodakarensis and requirement of tRNA m2G10/m22G10 methyltransferase (archaeal Trm11) for survival at high temperatures