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Literature summary extracted from

  • Akhtar, M.K.; Vijay, D.; Umbreen, S.; McLean, C.J.; Cai, Y.; Campopiano, D.J.; Loake, G.J.
    Hydrogen peroxide-based fluorometric assay for real-time monitoring of SAM-dependent methyltransferases (2018), Front. Bioeng. Biotechnol., 6, 146 .
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

EC Number Cloned (Comment) Organism
2.1.1.273 recombinant expression in Escherichia coli strain BL21(DE3) Clarkia breweri
2.1.1.274 recombinant expression in Escherichia coli strain BL21(DE3) Clarkia breweri

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
2.1.1.273 0.0068
-
 salicylate pH 7.5, 30°C Clarkia breweri
2.1.1.274 0.0068
-
 salicylate pH 7.5, 30°C Clarkia breweri

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
2.1.1.273 KCl activates at 1 mM Clarkia breweri
2.1.1.274 KCl activates at 1 mM Clarkia breweri

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
2.1.1.273 S-adenosyl-L-methionine + salicylate Clarkia breweri
-
 methyl salicylate + S-adenosyl-L-homocysteine
-
 ?
2.1.1.274 S-adenosyl-L-methionine + salicylate Clarkia breweri
-
 methyl salicylate + S-adenosyl-L-homocysteine
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
2.1.1.273 Clarkia breweri Q9SPV4 i.e. Eucharidium breweri
-
2.1.1.274 Clarkia breweri Q9SPV4 i.e. Eucharidium breweri
-

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2.1.1.273 additional information development and evaluation of an enzyme-coupled assay for monitoring methyltransferase activity, overview. Since S-adenosyl-L-homocysteine is a key by-product of reactions catalyzed by S-adenosyl methionine-dependent methyltransferases, the coupling enzymes are used to assess the activities of EcoRI methyltransferase and a salicylic acid methyltransferase from Clarkia breweri in the presence of S-adenosyl methionine. In the case of the salicylic acid methyltransferase, detectable activity is observed for several substrates including salicylic acid, benzoic acid, 3-hydroxybenzoic acid, and vanillic acid, substrate specificity, overview. Additionally, the de novo synthesis of the relatively expensive and unstable cosubstrate, S-adenosyl methionine, catalyzed by methionine adenosyltransferase can be incorporated within the assay. The assay offers a high level of sensitivity that permits continuous and reliable monitoring of methyltransferase activities. The assay enzymes, 5'-methylthioadenosine/S-adenosylhomocysteine nucleosidase (Mtn), xanthine oxidase (XOD), and horse radish peroxidase (HRP), are able to operate in a tandem manner to generate a fluorescence signal in the presence of SAH, the key by-product of reactions catalyzed by SAM-dependent methyltransferases. Poor or no substrates are acetate, propanoate, butyrate, 4-hydroxybenzoate, jasmonate, cinnamate, coumarate, and caffeate Clarkia breweri ?
-
-
2.1.1.273 S-adenosyl-L-methionine + 3-hydroxybenzoate 26% activity compared to salicylate Clarkia breweri methyl 3-hydroxybenzoate + S-adenosyl-L-homocysteine
-
 ?
2.1.1.273 S-adenosyl-L-methionine + benzoate 96% activity compared to salicylate Clarkia breweri methyl benzoate + S-adenosyl-L-homocysteine
-
 ?
2.1.1.273 S-adenosyl-L-methionine + salicylate
-
 Clarkia breweri methyl salicylate + S-adenosyl-L-homocysteine
-
 ?
2.1.1.273 S-adenosyl-L-methionine + salicylate best substrate Clarkia breweri methyl salicylate + S-adenosyl-L-homocysteine
-
 ?
2.1.1.273 S-adenosyl-L-methionine + vanillate 12% activity compared to salicylate Clarkia breweri methyl vanillate + S-adenosyl-L-homocysteine
-
 ?
2.1.1.274 additional information development and evaluation of an enzyme-coupled assay for monitoring methyltransferase activity, overview. Since S-adenosyl-L-homocysteine is a key by-product of reactions catalyzed by S-adenosyl methionine-dependent methyltransferases, the coupling enzymes are used to assess the activities of EcoRI methyltransferase and a salicylic acid methyltransferase from Clarkia breweri in the presence of S-adenosyl methionine. In the case of the salicylic acid methyltransferase, detectable activity is observed for several substrates including salicylic acid, benzoic acid, 3-hydroxybenzoic acid, and vanillic acid, substrate specificity, overview. Additionally, the de novo synthesis of the relatively expensive and unstable cosubstrate, S-adenosyl methionine, catalyzed by methionine adenosyltransferase can be incorporated within the assay. The assay offers a high level of sensitivity that permits continuous and reliable monitoring of methyltransferase activities. The assay enzymes, 5'-methylthioadenosine/S-adenosylhomocysteine nucleosidase (Mtn), xanthine oxidase (XOD), and horse radish peroxidase (HRP), are able to operate in a tandem manner to generate a fluorescence signal in the presence of SAH, the key by-product of reactions catalyzed by SAM-dependent methyltransferases. Poor or no substrates are acetate, propanoate, butyrate, 4-hydroxybenzoate, jasmonate, cinnamate, coumarate, and caffeate Clarkia breweri ?
-
-
2.1.1.274 S-adenosyl-L-methionine + 3-hydroxybenzoate 26% activity compared to salicylate Clarkia breweri methyl 3-hydroxybenzoate + S-adenosyl-L-homocysteine
-
 ?
2.1.1.274 S-adenosyl-L-methionine + benzoate 96% activity compared to salicylate Clarkia breweri methyl benzoate + S-adenosyl-L-homocysteine
-
 ?
2.1.1.274 S-adenosyl-L-methionine + salicylate
-
 Clarkia breweri methyl salicylate + S-adenosyl-L-homocysteine
-
 ?
2.1.1.274 S-adenosyl-L-methionine + salicylate best substrate Clarkia breweri methyl salicylate + S-adenosyl-L-homocysteine
-
 ?
2.1.1.274 S-adenosyl-L-methionine + vanillate 12% activity compared to salicylate Clarkia breweri methyl vanillate + S-adenosyl-L-homocysteine
-
 ?

Synonyms

EC Number Synonyms Comment Organism
2.1.1.273 More see also EC 2.1.1.274 Clarkia breweri
2.1.1.273 SA MTase
-
 Clarkia breweri
2.1.1.273 salicylic acid methyltransferase
-
 Clarkia breweri
2.1.1.274 More see also EC 2.1.1.273 Clarkia breweri
2.1.1.274 SA MTase
-
 Clarkia breweri
2.1.1.274 salicylic acid methyltransferase
-
 Clarkia breweri

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
2.1.1.273 30
-
 assay at Clarkia breweri
2.1.1.274 30
-
 assay at Clarkia breweri

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
2.1.1.273 0.0163
-
 salicylate pH 7.5, 30°C Clarkia breweri
2.1.1.274 0.0163
-
 salicylate pH 7.5, 30°C Clarkia breweri

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
2.1.1.273 7.5
-
 assay at Clarkia breweri
2.1.1.274 7.5
-
 assay at Clarkia breweri

Cofactor

EC Number Cofactor Comment Organism Structure
2.1.1.273 S-adenosyl-L-methionine
-
 Clarkia breweri
2.1.1.274 S-adenosyl-L-methionine
-
 Clarkia breweri

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
2.1.1.273 2.4
-
 salicylate pH 7.5, 30°C Clarkia breweri
2.1.1.274 2.4
-
 salicylate pH 7.5, 30°C Clarkia breweri