Literature summary for 2.1.1.204 extracted from

Johannsson, S.; Neumann, P.; Wulf, A.; Welp, L.M.; Gerber, H.D.; Krull, M.; Diederichsen, U.; Urlaub, H.; Ficner, R.
Structural insights into the stimulation of S. pombe Dnmt2 catalytic efficiency by the tRNA nucleoside queuosine (2018), Sci. Rep., 8, 8880 .

Search for more literature for 2.1.1.204

Cloned(Commentary)

Cloned (Comment)	Organism
recombinant expression of GST-tagged enzyme in Escherichia coli strain BL21(DE3)	Schizosaccharomyces pombe

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified recombinant enzyme Dnmt2, sitting-drop vapor diffusion technique, mixing of equal volumes of 2 mg/ml protein solution and crystallization buffer containing 50 mM MES, pH 6.5, 1% w/v PEG 4000, and 4 mM MgCl2, 4°C, 5-7 days, X-ray diffraction structure determination and analysis at 1.7 A resolution, molecular replacement using the structure of Entamoebae histolytica Dnmt2 homologue EhMeth (PBD ID 3QV2), modelling	Schizosaccharomyces pombe

Crystallization (Comment)

Organism

purified recombinant enzyme Dnmt2, sitting-drop vapor diffusion technique, mixing of equal volumes of 2 mg/ml protein solution and crystallization buffer containing 50 mM MES, pH 6.5, 1% w/v PEG 4000, and 4 mM MgCl2, 4°C, 5-7 days, X-ray diffraction structure determination and analysis at 1.7 A resolution, molecular replacement using the structure of Entamoebae histolytica Dnmt2 homologue EhMeth (PBD ID 3QV2), modelling

Schizosaccharomyces pombe

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism
additional information	-	additional information	Q34tRNAAsp exhibits an only slightly increased affinity for Dnmt2 in comparison to unmodified G34tRNA. Kinetic analysis, overview. Quantitative analysis of Dnmt2 tRNA complex formation	Schizosaccharomyces pombe
0.00688	-	cytosine38 in Q34tRNAAsp	pH 7.5, temperature not specified in the publication	Schizosaccharomyces pombe
0.01559	-	cytosine38 in G34tRNAAsp	pH 7.5, temperature not specified in the publication	Schizosaccharomyces pombe

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.
S-adenosyl-L-methionine + cytosine38 in G34tRNAAsp	Schizosaccharomyces pombe	-	S-adenosyl-L-homocysteine + 5-methylcytosine38 in G34tRNAAsp	-	?
S-adenosyl-L-methionine + cytosine38 in G34tRNAAsp	Schizosaccharomyces pombe ATCC 24843	-	S-adenosyl-L-homocysteine + 5-methylcytosine38 in G34tRNAAsp	-	?
S-adenosyl-L-methionine + cytosine38 in G34tRNAAsp	Schizosaccharomyces pombe 972	-	S-adenosyl-L-homocysteine + 5-methylcytosine38 in G34tRNAAsp	-	?
S-adenosyl-L-methionine + cytosine38 in Q34tRNAAsp	Schizosaccharomyces pombe	preferred substrate	S-adenosyl-L-homocysteine + 5-methylcytosine38 in Q34tRNAAsp	-	?
S-adenosyl-L-methionine + cytosine38 in Q34tRNAAsp	Schizosaccharomyces pombe ATCC 24843	preferred substrate	S-adenosyl-L-homocysteine + 5-methylcytosine38 in Q34tRNAAsp	-	?
S-adenosyl-L-methionine + cytosine38 in Q34tRNAAsp	Schizosaccharomyces pombe 972	preferred substrate	S-adenosyl-L-homocysteine + 5-methylcytosine38 in Q34tRNAAsp	-	?

Organism

Organism	UniProt	Comment	Textmining
Schizosaccharomyces pombe	P40999	-	-
Schizosaccharomyces pombe 972	P40999	-	-
Schizosaccharomyces pombe ATCC 24843	P40999	-	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant GST-tagged enzyme from Escherichia coli strain BL21(DE3) by ultracentrifugation, glutathione affinity chromatography, and tag cleavage through PreScission protease, followed by heparin affinity chromatography, gel filtration, and ultrafiltration	Schizosaccharomyces pombe

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
additional information	Q34tRNAAsp exhibits an only slightly increased affinity for Dnmt2 in comparison to unmodified G34tRNA	Schizosaccharomyces pombe	?	-	-
additional information	Q34tRNAAsp exhibits an only slightly increased affinity for Dnmt2 in comparison to unmodified G34tRNA	Schizosaccharomyces pombe ATCC 24843	?	-	-
additional information	Q34tRNAAsp exhibits an only slightly increased affinity for Dnmt2 in comparison to unmodified G34tRNA	Schizosaccharomyces pombe 972	?	-	-
S-adenosyl-L-methionine + cytosine38 in G34tRNAAsp	-	Schizosaccharomyces pombe	S-adenosyl-L-homocysteine + 5-methylcytosine38 in G34tRNAAsp	-	?
S-adenosyl-L-methionine + cytosine38 in G34tRNAAsp	-	Schizosaccharomyces pombe ATCC 24843	S-adenosyl-L-homocysteine + 5-methylcytosine38 in G34tRNAAsp	-	?
S-adenosyl-L-methionine + cytosine38 in G34tRNAAsp	-	Schizosaccharomyces pombe 972	S-adenosyl-L-homocysteine + 5-methylcytosine38 in G34tRNAAsp	-	?
S-adenosyl-L-methionine + cytosine38 in Q34tRNAAsp	preferred substrate	Schizosaccharomyces pombe	S-adenosyl-L-homocysteine + 5-methylcytosine38 in Q34tRNAAsp	-	?
S-adenosyl-L-methionine + cytosine38 in Q34tRNAAsp	preferred substrate	Schizosaccharomyces pombe ATCC 24843	S-adenosyl-L-homocysteine + 5-methylcytosine38 in Q34tRNAAsp	-	?
S-adenosyl-L-methionine + cytosine38 in Q34tRNAAsp	preferred substrate	Schizosaccharomyces pombe 972	S-adenosyl-L-homocysteine + 5-methylcytosine38 in Q34tRNAAsp	-	?

Synonyms

Synonyms	Comment	Organism
Dnmt2	-	Schizosaccharomyces pombe
spDnmt2	-	Schizosaccharomyces pombe

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
7.5	-	assay at	Schizosaccharomyces pombe

Cofactor

Cofactor	Comment	Organism	Structure
S-adenosyl-L-methionine	-	Schizosaccharomyces pombe

General Information

General Information	Comment	Organism
additional information	enzyme Dnmt2 from Schizosaccharomyces pombe contains the entire active site loop. The interaction with tRNA is analyzed by means of mass spectrometry using UV cross-linked Dnmt2-tRNA complex. Cross-link data and computational docking of Dnmt2 and tRNAAsp reveal Q34 positioned adjacent to the S-adenosylmethionine occupies the active site, suggesting that the observed increase of Dnmt2 catalytic efficiency by queuine originates from optimal positioning of the substrate molecules and residues relevant for methyl transfer. Observation of displacement of the cytosine from the codon-anticodon helix and distortion of the latter, hinting to a role of the Q-modification in translational accuracy. Analysis of spDnmt2 electrostatic surface potentials unveils predominantly positively charged surface around the SAH indicating possible interaction with the tRNA phosphate backbone. But structure exposes a strongly negatively charged cavity in close proximity to the sulfur of S-adenosyl-L-homocysteine formed by the conserved catalytic residue Glu121	Schizosaccharomyces pombe
physiological function	enzyme Dnmt2 methylates cytosine at position 38 of tRNAAsp. A correlation between the presence of the hypermodified nucleoside queuosine (Q) at position 34 of tRNAAsp and the Dnmt2 dependent C38 methylation has recently been found in vivo for Schizosaccharomcyces pombe. Dnmt2 shows an increase for in vitro transcribed tRNAAsp containing Q34 compared to the unmodified substrate, structural basis for the Q-dependency, overview. The C38 methylation of tRNAAsp in Schizosaccharomcyces pombe depends on the presence of queuosine (Q), a hypermodified nucleoside at position 34 of tRNAAsp, tRNAAsn, tRNATyr, and tRNAHis10	Schizosaccharomyces pombe