Cloned (Comment) | Organism |
---|---|
Clarkia breweri SAMT cloned into expression vector pET28a(+) and construct transformed into Escherichia coli BL21(DE3) cells | Clarkia breweri |
Crystallization (Comment) | Organism |
---|---|
overall structure of SAMT monomer consists of a globular domain containing the extended beta-sheet characteristic of other SAM-dependent methyltransferases and a unique alpha-helical cap that forms the top one-third of the active site cavity | Clarkia breweri |
SAMT crystallized from ammonium sulfate solution, model spanned the entire 359 residues of full-length Clarkia SAMT | Clarkia breweri |
Protein Variants | Comment | Organism |
---|---|---|
Y147S | reduced methylation of benzoic acid and 3-hydroxybenzoic acid but slightly increased activity towards jasmonic acid | Clarkia breweri |
Y147S/M150H | significant increase in the ability to turn over jasmonic acid and vanillic acid | Clarkia breweri |
Y147S/M150H/F347Y | slightly increased activity towards short-chain carboxylic acids and jasmonic acid | Clarkia breweri |
Y147S/M150H/F347Y/N349I | highly increased activity towards short-chain carboxylic acids and aromatic acids like hydroxybenzoic acids, vanillic acid, jasmonic acid, cinnamic acid, 4-coumaric acid and caffeic acid, broadest substrate spectrum | Clarkia breweri |
Y147S/M150H/I225Q/F347Y | greatest specific activities against 3-hydroxybenzoic acid, vanillic acid and jasmonic acid but reduced activity towards salicylic acid | Clarkia breweri |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.023 | - |
salicylate | wild type enzyme, pH 7.5, 25°C | Clarkia breweri |
Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|
41000 | - |
SDS-PAGE | Clarkia breweri |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Clarkia breweri | Q9SPV4 | - |
- |
Purification (Comment) | Organism |
---|---|
N-terminal polyhistidine-tagged SAMT protein purified by Ni2+ affinity chromatography and gel filtration chromatography using a Superdex-75 column | Clarkia breweri |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
additional information | no measurable methylation of jasmonic acid by wild-type SAMT using concentrations up to 5 mM | Clarkia breweri | ? | - |
? | |
additional information | wild-type is also able to methylate benzoic acid with 48% activity compared to salicylate methylation | Clarkia breweri | ? | - |
? | |
S-adenosyl-L-methionine + 3-hydroxybenzoic acid | 17% activity of wild-type enzyme compared to salicylate methylation | Clarkia breweri | S-adenosyl-L-homocysteine + methyl 3-hydroxybenzoate | - |
? | |
S-adenosyl-L-methionine + jasmonic acid | Y147S/M150H double mutant and Y147S/M150H/F347Y triple mutant are able to turn over jasmonic acid, while preserving substantial salicylate methylating activity | Clarkia breweri | S-adenosyl-L-homocysteine + methyl jasmonate | - |
? | |
S-adenosyl-L-methionine + salicylate | - |
Clarkia breweri | S-adenosyl-L-homocysteine + methyl salicylate | - |
? | |
S-adenosyl-L-methionine + vanillic acid | 5.1% activity of wild-type enzyme compared to salicylate methylation | Clarkia breweri | S-adenosyl-L-homocysteine + methyl 4-hydroxy-3-methoxybenzoate | - |
? |
Subunits | Comment | Organism |
---|---|---|
homodimer | 2 * 41000 | Clarkia breweri |
Synonyms | Comment | Organism |
---|---|---|
SA carboxyl methyltransferase | - |
Clarkia breweri |
SAMT | - |
Clarkia breweri |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.092 | - |
salicylate | wild type enzyme, pH 7.5, 25°C | Clarkia breweri |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
4 | - |
salicylate | wild type enzyme, pH 7.5, 25°C | Clarkia breweri |