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2.1.1.202: multisite-specific tRNA:(cytosine-C5)-methyltransferase

This is an abbreviated version!
For detailed information about multisite-specific tRNA:(cytosine-C5)-methyltransferase, go to the full flat file.

Word Map on EC 2.1.1.202

Reaction

S-adenosyl-L-methionine
+
cytosine34 in tRNA precursor
=
S-adenosyl-L-homocysteine
 +
5-methylcytosine34 in tRNA precursor

Synonyms

5-methylcytosine RNA methyltransferase, broad-spectrum methyltransferase, cytosine-5 RNA methyltransferase, EC 2.1.1.29, HNSun6, M5C RNA methyltransferase, MJ0026, multisite-specific tRNA:m5C-methyltransferase, Ncl1p, NOP2/Sun RNA methyltransferase 2, NSUN2, NSUN6, PhNSUN6, RNA (cytosine-5)-methyltransferase, RNA:m5C methyltransferase, RNA:m5C MTase, TRDMT1, TRM4, TRM4A, TRM4B, Trm4p, tRNA (cytosine72-C5)-methyltransferase, tRNA-specific methyltransferase 4B, tRNA:m5C-methyltransferase, tRNA:m5C72 MTase

ECTree

     2 Transferases
         2.1 Transferring one-carbon groups
             2.1.1 Methyltransferases
                2.1.1.202 multisite-specific tRNA:(cytosine-C5)-methyltransferase

Crystallization

Crystallization on EC 2.1.1.202 - multisite-specific tRNA:(cytosine-C5)-methyltransferase

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CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
crystal structure of NSUN6 with target RNA showing the arrangement and key contacts between the amino acids in the active site and the target cytosine (C72) in RNA (PDB ID 5WWS)
purified recombinant His6-tagged enzyme, hanging drop vapor diffusion method, mixing of 0.001 ml of 4.8 mg/ml protein in 20 mM Tris-HCl, pH 7.0, 300 mM NaCl, 5 mM MgCl2, and 5 mM DTT, with 0.001 ml of reservoir solution consisting of 2% dioxane, 0.1 M bicine, pH 8.6, 18% w/v PEG 8000, and 10 mM yttrium(III) chloride hexahydrate, 16°C. For the hNSun6/tRNA complex, solutions are prepared with 0.075 mM MhNSun6 and 0.075 mM MtRNA, for the hNSun6/tRNA/SAM and hNSun6/tRNA/SFG complexes, hNSun6 is mixed with tRNA at a 1:1 molar ratio, adding either 1 mM SAM or SFG, crystals are obtained by mixing 0.001 ml of this solution with 0.001 ml of reservoir solution containing 0.1 M SPG, pH 8.0-10.0, 24-28% w/v PEG 1500, and 50 mM NaF, X-ray diffraction structure determination and analysis. Of different human tRNACys and tRNAThr isoacceptors and their variants, only tRNACys(GCA)-G2A:C71U can be cocrystallized with hNSun6, molecular replacement using the MTase domain structure of Pyrococcus horikoshii PH0851 (PDB ID: 2YXL) and the PUA domain structure of Pyrococcus horikoshii archaeosine tRNA-guanine transglycosylase (PDB ID: 1J2B) as starting models
Trm4 free and in complex with sinefungin, hanging-drop vapor diffusion, 0.001 ml of protein solution containing 10.8 mg/ml protein with 0.001 ml of crystallization solution containing 100 mM Tris-HCl, pH 7.4, 200 mM MgCl2, 25% PEG 3350, and 6% v/v isopropanol for MJ0026 alone, and 20 100 mM Tris-HCl, pH 7.4, 200 mM MgCl2, 25% PEG 3350, 3 mM sinefungin, and 10 mM cytidine for Mj0026 with sinefungin, cryoprotectant 20% glycerol and 20% PEG 400, respectively, X-ray diffraction structure determination and analysis at 1.27 A and 2.3 A resolutions, respectively
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