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Literature summary extracted from

  • Chen, H.; Yamase, H.; Murakami, K.; Chang, C.W.; Zhao, L.; Zhao, Z.; Liu, H.W.
    Expression, purification, and characterization of two N,N-dimethyltransferases, tylM1 and desVI, involved in the biosynthesis of mycaminose and desosamine (2002), Biochemistry, 41, 9165-9183.
    View publication on PubMed

Cloned(Commentary)

EC Number Cloned (Comment) Organism
2.1.1.234 expression in Escherichia coli Streptomyces venezuelae
2.1.1.235 expression in Escherichia coli. Heterologous expression of the tylM1 gene in the desVI deletion mutant of Streptomyces venezuelae. TylM1 is a competent substitute for DesVI Streptomyces fradiae

General Stability

EC Number General Stability Organism
2.1.1.234 inclusion of 0.1 mM S-adenosyl-L-methionine, 0.1 mM dithiothreitol, and 10% glycerol in in buffers is crucial to prevent TylM1 from precipitating during the purification Streptomyces venezuelae
2.1.1.235 concentrated TylM1 protein is stable and can endure repeated freeze/thaw cycles without losing noticeable activity Streptomyces fradiae
2.1.1.235 enzyme Streptomyces fradiae
2.1.1.235 inclusion of 0.1 mM S-adenosyl-L-methioninet in buffers is crucial to prevent TylM1 from precipitating during the purification Streptomyces fradiae
2.1.1.235 TylM1 is unstable at low concentrations, requiring the addition of bovine serum albumin (1 mg/mL) to the assay buffer during kinetic studies of the Streptomyces fradiae

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
2.1.1.234 0.2766
-
 dTDP-3-amino-3,6-dideoxy-alpha-D-glucopyranose pH 7.5, 24°C, cosubstrate: 3.0 mM S-adenosyl-L-methionine Streptomyces venezuelae
2.1.1.234 0.3074
-
 dTDP-3-amino-3,4,6-trideoxy-alpha-D-glucose pH 7.5, 24°C, cosubstrate: 3.0 mM S-adenosyl-L-methionine Streptomyces venezuelae
2.1.1.235 0.0468
-
 dTDP-3-methylamino-3,6-dideoxy-alpha-D-glucopyranose pH 7.5, 24°C Streptomyces fradiae
2.1.1.235 0.0594
-
 dTDP-3-amino-3,6-dideoxy-alpha-D-glucopyranose pH 7.5, 24°C Streptomyces fradiae
2.1.1.235 0.0989
-
 S-adenosyl-L-methionine pH 7.5, 24°C, cosubstrate: 1.28 mM dTDP-3-amino-3,6-dideoxy-alpha-D-glucopyranose Streptomyces fradiae
2.1.1.235 0.117
-
 S-adenosyl-L-methionine pH 7.5, 24°C, cosubstrate: dTDP-3-methylamino-3,6-dideoxy-alpha-D-glucopyranose Streptomyces fradiae
2.1.1.235 0.1184
-
 dTDP-3-amino-3,6-dideoxy-alpha-D-glucopyranose pH 7.5, 24°C Streptomyces fradiae

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
2.1.1.235 additional information TylM1 is not a Mg2+-dependent methyltransferase Streptomyces fradiae

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
2.1.1.234 28000
-
 2 * 28000, SDS-PAGE Streptomyces venezuelae
2.1.1.234 47400
-
 gel filtration Streptomyces venezuelae
2.1.1.235 27000
-
 2 * 27000, SDS-PAGE Streptomyces fradiae
2.1.1.235 27427
-
 2 * 27427, calculated from sequence Streptomyces fradiae
2.1.1.235 55200
-
 gel filtration Streptomyces fradiae

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
2.1.1.234 2 S-adenosyl-L-methionine + dTDP-3-amino-4,6-dideoxy-alpha-D-glucopyranose Streptomyces venezuelae the enzyme is involved in the biosynthesis of desosamine 2 S-adenosyl-L-homocysteine + dTDP-3-dimethylamino-4,6-dideoxy-alpha-D-glucopyranose
-
 ?
2.1.1.235 2 S-adenosyl-L-methionine + dTDP-3-amino-3,6-dideoxy-alpha-D-glucopyranose Streptomyces fradiae the enzyme is involved in the biosynthesis of mycaminose, an essential structural component of the macrolide antibiotic tylosin poduced by Streptomyces fradiae 2 S-adenosyl-L-homocysteine + dTDP-3-dimethylamino-3,6-dideoxy-alpha-D-glucopyranose
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
2.1.1.234 Streptomyces venezuelae Q9ZGH6
-
-
2.1.1.235 Streptomyces fradiae P95748
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
2.1.1.234
-
 Streptomyces venezuelae
2.1.1.235 inclusion of 0.1 mM AdoMet in buffers is crucial to prevent TylM1 from precipitating during the purification Streptomyces fradiae

Storage Stability

EC Number Storage Stability Organism
2.1.1.235 -80 °C concentrated TylM1 protein is stable for at least 2 years Streptomyces fradiae

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2.1.1.234 2 S-adenosyl-L-methionine + dTDP-3-amino-3,6-dideoxy-alpha-D-glucopyranose the kcat/KM for dTDP-3-amino-3,6-dideoxy-alpha-D-glucopyranose is about 20fold lower than kcat/Km for the natural substrated TDP-3-amino-4,6-dideoxy-alpha-D-glucopyranose Streptomyces venezuelae 2 S-adenosyl-L-homocysteine + dTDP-3-dimethylamino-3,6-dideoxy-alpha-D-glucopyranose
-
 ?
2.1.1.234 2 S-adenosyl-L-methionine + dTDP-3-amino-4,6-dideoxy-alpha-D-glucopyranose the enzyme is involved in the biosynthesis of desosamine Streptomyces venezuelae 2 S-adenosyl-L-homocysteine + dTDP-3-dimethylamino-4,6-dideoxy-alpha-D-glucopyranose
-
 ?
2.1.1.234 2 S-adenosyl-L-methionine + TDP-3-amino-4,6-dideoxy-alpha-D-glucopyranose i.e. TDP-alpha-D-desosamine. When the reaction is quenched at an earlier time, the monomethylated species can also be detected Streptomyces venezuelae 2 S-adenosyl-L-homocysteine + dTDP-3-dimethylamino-4,6-dideoxy-alpha-D-glucopyranose
-
 ?
2.1.1.235 2 S-adenosyl-L-methionine + dTDP-3-amino-3,4,6-trideoxy-alpha-xylo-hexopyranose kcat/KM is about 3fold lower than kcat/Km for the natural substrated TDP-3-amino-3,6-dideoxy-alpha-D-glucopyranose Streptomyces fradiae 2 S-adenosyl-L-homocysteine + dTDP-3-dimethylamino-3,4,6-trideoxy-alpha-xylo-hexopyranose i.e. TDP-alpha-D-desosamine ?
2.1.1.235 2 S-adenosyl-L-methionine + dTDP-3-amino-3,6-dideoxy-alpha-D-glucopyranose the enzyme is involved in the biosynthesis of mycaminose, an essential structural component of the macrolide antibiotic tylosin poduced by Streptomyces fradiae Streptomyces fradiae 2 S-adenosyl-L-homocysteine + dTDP-3-dimethylamino-3,6-dideoxy-alpha-D-glucopyranose
-
 ?
2.1.1.235 2 S-adenosyl-L-methionine + dTDP-3-amino-3,6-dideoxy-alpha-D-glucopyranose TylM1 has relaxed specificity toward its sugar substrate. The kcat/KM for dTDP-3-amino-4,6-dideoxy-alpha-D-glucopyranose is about 3fold lower than kcat/Km for the natural substrated TDP-3-amino-3,6-dideoxy-alpha-D-glucopyranose Streptomyces fradiae 2 S-adenosyl-L-homocysteine + dTDP-3-dimethylamino-3,6-dideoxy-alpha-D-glucopyranose
-
 ?
2.1.1.235 S-adenosyl-L-methionine + dTDP-3-methylamino-3,6-dideoxy-alpha-D-glucopyranose chemically prepared monomethylated compound is a substrate for TylM1 and could be swiftly converted by TylM1 to the dimethylated product. TylM1 catalyzes an N,N-dimethylation reaction by way of a monomethylated intermediate. Since the monomethylated amino group is intrinsically a better nucleophile than the unsubstituted amino group, the reaction rate of this SN2-type methyl transfer reaction is expected to be higher for the second half reaction than for the first methylation reaction Streptomyces fradiae S-adenosyl-L-homocysteine + dTDP-3-dimethylamino-3,6-dideoxy-alpha-D-glucopyranose
-
 ?

Subunits

EC Number Subunits Comment Organism
2.1.1.234 homodimer 2 * 28000, SDS-PAGE Streptomyces venezuelae
2.1.1.234 homodimer 2 * 28110, calculated from sequence Streptomyces venezuelae
2.1.1.235 homodimer 2 * 27000, SDS-PAGE Streptomyces fradiae
2.1.1.235 homodimer 2 * 27427, calculated from sequence Streptomyces fradiae

Synonyms

EC Number Synonyms Comment Organism
2.1.1.234 DesVI
-
 Streptomyces venezuelae
2.1.1.235 TylM1
-
 Streptomyces fradiae

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
2.1.1.234 24
-
 assay at Streptomyces venezuelae
2.1.1.235 24
-
 assay at Streptomyces fradiae

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
2.1.1.234 0.07
-
 dTDP-3-amino-3,6-dideoxy-alpha-D-glucopyranose pH 7.5, 24°C, cosubstrate: 3.0 mM S-adenosyl-L-methionine Streptomyces venezuelae
2.1.1.234 1.533
-
 dTDP-3-amino-3,4,6-trideoxy-alpha-D-glucose pH 7.5, 24°C, cosubstrate: 3.0 mM S-adenosyl-L-methionine Streptomyces venezuelae
2.1.1.235 0.09
-
 S-adenosyl-L-methionine pH 7.5, 24°C, cosubstrate: 1.28 mM dTDP-3-amino-3,6-dideoxy-alpha-D-glucopyranose Streptomyces fradiae
2.1.1.235 0.12
-
 dTDP-3-amino-3,6-dideoxy-alpha-D-glucopyranose pH 7.5, 24°C Streptomyces fradiae
2.1.1.235 0.165
-
 dTDP-3-amino-3,6-dideoxy-alpha-D-glucopyranose pH 7.5, 24°C Streptomyces fradiae
2.1.1.235 0.54
-
 dTDP-3-methylamino-3,6-dideoxy-alpha-D-glucopyranose pH 7.5, 24°C Streptomyces fradiae
2.1.1.235 0.86
-
 S-adenosyl-L-methionine pH 7.5, 24°C, cosubstrate: 0.89 mM dTDP-3-methylamino-3,6-dideoxy-alpha-D-glucopyranose Streptomyces fradiae

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
2.1.1.234 7.5
-
 assay at Streptomyces venezuelae
2.1.1.235 7.5
-
 assay at Streptomyces fradiae

Cofactor

EC Number Cofactor Comment Organism Structure
2.1.1.234 S-adenosyl-L-methionine
-
 Streptomyces venezuelae
2.1.1.235 S-adenosyl-L-methionine
-
 Streptomyces fradiae

General Information

EC Number General Information Comment Organism
2.1.1.234 physiological function the enzyme catalyzes the N,N-dimethylation step in the biosynthesis of desosamine Streptomyces venezuelae
2.1.1.235 physiological function the enzyme catalyzes the N,N-dimethylation step in the biosynthesis of mycaminose. Mycaminose is an aminohexose found in several macrolide antibiotics. The sugar contains a C-3 N,N-dimethylamino group which confers the biological activity of these unusual sugar Streptomyces fradiae

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
2.1.1.234 0.25
-
 dTDP-3-amino-3,6-dideoxy-alpha-D-glucopyranose pH 7.5, 24°C, cosubstrate: 3.0 mM S-adenosyl-L-methionine Streptomyces venezuelae
2.1.1.234 5
-
 dTDP-3-amino-3,4,6-trideoxy-alpha-D-glucose pH 7.5, 24°C, cosubstrate: 3.0 mM S-adenosyl-L-methionine Streptomyces venezuelae
2.1.1.235 0.91
-
 S-adenosyl-L-methionine pH 7.5, 24°C, cosubstrate: 1.28 mM dTDP-3-amino-3,6-dideoxy-alpha-D-glucopyranose Streptomyces fradiae
2.1.1.235 1
-
 dTDP-3-amino-3,6-dideoxy-alpha-D-glucopyranose pH 7.5, 24°C Streptomyces fradiae
2.1.1.235 2.78
-
 dTDP-3-amino-3,6-dideoxy-alpha-D-glucopyranose pH 7.5, 24°C Streptomyces fradiae
2.1.1.235 7.35
-
 S-adenosyl-L-methionine pH 7.5, 24°C, cosubstrate: dTDP-3-methylamino-3,6-dideoxy-alpha-D-glucopyranose Streptomyces fradiae
2.1.1.235 11.5
-
 dTDP-3-methylamino-3,6-dideoxy-alpha-D-glucopyranose pH 7.5, 24°C Streptomyces fradiae