Cloned (Comment) | Organism |
---|---|
genes GCD10 and GCD14, recombinant co-expression of wild-type and selenomethionine-labeled His-tagged subunits in Escherichia coli | Saccharomyces cerevisiae |
Crystallization (Comment) | Organism |
---|---|
purified recombinant wild-type and selenomethionine-labeled holoenzyme in apoform and complexed with S-adenosyl-L-methionine (SAM), mixing of 15 mg/ml protein in 20 mM Tris-HCl, pH 8.0, 300 mM NaCl, and 5 mM DTT, with a three-fold molecular excess of S-adenosyl-L-methionine, sitting drop vapour diffusion method, with the mother liquor containing 0.1 M HEPES, PH 7.5, 2% v/v 2-methyl-2,4-pentanediol, 10% w/v PEG 6000, 3 days X-ray diffraction structure determination and analysis | Saccharomyces cerevisiae |
Protein Variants | Comment | Organism |
---|---|---|
additional information | the human homologue of the yeast tRNA m1A58 methyltransferase is identified through amino acid sequence identity and complementation of the yeast temperature-sensitive TRM6 andTRM61 mutant phenotypes27. When co-expressed in yeast, the Homo sapiens TRM6-TRM61 catalyzes the in vitro methyl transfer reaction for both the yeast initiator tRNAi Met and human tRNA3Lys27 | Saccharomyces cerevisiae |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | thermodynamics | Saccharomyces cerevisiae |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
S-adenosyl-L-methionine + adenine58 in tRNA | Saccharomyces cerevisiae | - |
S-adenosyl-L-homocysteine + N1-methyladenine58 in tRNA | - |
? | |
S-adenosyl-L-methionine + adenine58 in tRNA | Saccharomyces cerevisiae ATCC 204508 | - |
S-adenosyl-L-homocysteine + N1-methyladenine58 in tRNA | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Saccharomyces cerevisiae | P41814 AND P46959 | subunits Trm6 and Trm61 | - |
Saccharomyces cerevisiae ATCC 204508 | P41814 AND P46959 | subunits Trm6 and Trm61 | - |
Purification (Comment) | Organism |
---|---|
recombinant wild-type and selenomethionine-labeled His-tagged subunits Trm6 and Trm61 from Escherichia coli by nickel affinity chromatography and gel filtration | Saccharomyces cerevisiae |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
S-adenosyl-L-methionine + adenine58 in tRNA | - |
Saccharomyces cerevisiae | S-adenosyl-L-homocysteine + N1-methyladenine58 in tRNA | - |
? | |
S-adenosyl-L-methionine + adenine58 in tRNA | - |
Saccharomyces cerevisiae ATCC 204508 | S-adenosyl-L-homocysteine + N1-methyladenine58 in tRNA | - |
? |
Subunits | Comment | Organism |
---|---|---|
heterodimer | TRM6 and TRM61 form a compact complex via numerous hydrogen bonding and extensive hydrophobic interactions. The heterodimer interface of TRM6-TRM61 buries 3194 A2 of TRM6 and 3167 A2 of TRM61 solvent-accessible area, which represents about 17% and 16% of TRM6 and TRM61's total surface area, respectively. TRM6 mainly interacts with TRM61 through four major sites. Interaction analyses for sites A-C, detailed overview | Saccharomyces cerevisiae |
heterotetramer | two TRM6-TRM61 heterodimers assemble as a heterotetramer. A symmetric unit of the TRM6-TRM61 crystal contains one molecule of TRM6 and one molecule of TRM61, forming a 1:1 heterodimer. TRM6 and TRM61 form a 2:2 tetrameric heterocomplex, displaying an omega shape. Two symmetry-related TRM6-TRM61 heterodimer come together to form a central beta-barrel structure that consists of beta13 (TRM6), loop beta13/beta14 (TRM6), beta12(TRM61) and loop beta13/beta14 (TRM61). The top of the barrel contains a hydrophobic core, formed by residues Tyr422 (TRM6), Pro431 (TRM6), Met253 (TRM61), His354 (TRM61), and Tyr357 (TRM61) The center of the barrel is filled with numerous hydrophilic side-chains, including residues Glu416 (TRM6), Arg418 (TRM6), Arg420 (TRM6), Glu255 (TRM61), Gln257 (TRM61) and Arg259 (TRM61). The bottom of the barrel consists of a cage of four tyrosine residues. Modelling of the heterotetramer interface of TRM6-TRM61, overview. A TRM6-TRM61 heterotetramer constitutes two L-shaped tRNA binding regions. Structure comparison to the enzyme from Homo sapiens | Saccharomyces cerevisiae |
Synonyms | Comment | Organism |
---|---|---|
m1A58 tRNA methyl-transferase | - |
Saccharomyces cerevisiae |
TRM6-TRM61 holoenzyme | - |
Saccharomyces cerevisiae |
TrmI | - |
Saccharomyces cerevisiae |
tRNA m(1)A58 methyltransferase | - |
Saccharomyces cerevisiae |
two component m1A58 tRNA methyl-transferase | - |
Saccharomyces cerevisiae |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
8 | - |
assay at | Saccharomyces cerevisiae |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
S-adenosyl-L-methionine | SAM is situated in a cleft on the surface of the C-terminal Rossmann-fold domain of TRM61. The interaction between TRM61 and SAM can be divided into three parts in accordance to the moieties of SAM. For the adenine moiety, the side-chain atom OD1 of Asp168 makes a hydrogen bond with the nitrogen N6 of the adenine. Modeling of the SAM binding site and a possible adenine-binding pocket of TRM6-TRM61 | Saccharomyces cerevisiae |
General Information | Comment | Organism |
---|---|---|
evolution | TRM61 is the eukaryotic homologue of the bacterial and archaeal m1A58 tRNA methyl-transferase TrmI. Evolutionary relationship between TRM6 and TRM61, overview | Saccharomyces cerevisiae |
additional information | two TRM6-TRM61 heterodimers assemble as a heterotetramer. Both TRM6 and TRM61 subunits comprise an N-terminal beta-barrel domain linked to a C-terminal Rossmann-fold domain. TRM61 functions as the catalytic subunit, containing a methyl donor (SAM) binding pocket. TRM6 diverges from TRM61, lacking the conserved motifs used for binding SAM. TRM6 cooperates with TRM61 forming an L-shaped tRNA binding regions. Target tRNA recognition and catalytic mechanism of the two component m1A58 tRNA methyl-transferase, overview | Saccharomyces cerevisiae |
physiological function | the N1 methylation of adenine at position 58 (m1A58) of tRNA is an important post-transcriptional modification, which is vital for maintaining the stability of the initiator methionine tRNAiMet. Adenine at position 58 (A58) located in the T-loop is one of the most conserved nucleosides in tRNA. In eukaryotes, this modification is performed by the TRM6-TRM61 holoenzyme, molecular mechanism that underlies the cooperation of TRM6 and TRM61 in the methyl transfer reaction, overview | Saccharomyces cerevisiae |