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Information on EC 2.1.1.274 - salicylate 1-O-methyltransferase and Organism(s) Clarkia breweri and UniProt Accession Q9SPV4
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EC Tree
2.1.1.274 salicylate 1-O-methyltransferaseIUBMB Comments
The enzyme, which is found in flowering plants, also has the activity of EC 2.1.1.273, benzoate O-methyltransferase.
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The taxonomic range for the selected organisms is: Clarkia breweri
The enzyme appears in selected viruses and cellular organisms
The enzyme appears in selected viruses and cellular organisms
Reaction Schemes
Synonyms
samt1, cssamt, salicylic acid carboxyl methyltransferase, s-adenosyl-l-methionine salicylic acid carboxyl methyltransferase, sa methyltransferase, salicylic acid methyl transferase, ptsabath4, sa methyl transferase, pasabath2, sa carboxyl methyltransferase, 
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topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
S-adenosyl-L-methionine + salicylate = S-adenosyl-L-homocysteine + methyl salicylate
the enzyme, which is found in flowering plants, also has the activity of EC 2.1.1.273, benzoate O-methyltransferase
PATHWAY SOURCE
PATHWAYS
-
-, -
SYSTEMATIC NAME
IUBMB Comments
S-adenosyl-L-methionine:salicylate 1-O-methyltransferase
The enzyme, which is found in flowering plants, also has the activity of EC 2.1.1.273, benzoate O-methyltransferase.
SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
S-adenosyl-L-methionine + 3-hydroxybenzoate
methyl 3-hydroxybenzoate + S-adenosyl-L-homocysteine
26% activity compared to salicylate
-
-
?
S-adenosyl-L-methionine + 3-hydroxybenzoic acid
S-adenosyl-L-homocysteine + methyl 3-hydroxybenzoate
S-adenosyl-L-methionine + benzoate
methyl benzoate + S-adenosyl-L-homocysteine
96% activity compared to salicylate
-
-
?
S-adenosyl-L-methionine + cinnamic acid
S-adenosyl-L-homocysteine + methyl cinnamate
less than 2% relative activity at 1 mM methyl acceptor compared to activity with salicylate set at 100%
-
-
?
S-adenosyl-L-methionine + jasmonic acid
S-adenosyl-L-homocysteine + methyl jasmonate
Y147S/M150H double mutant and Y147S/M150H/F347Y triple mutant are able to turn over jasmonic acid, while preserving substantial salicylate methylating activity
-
-
?
S-adenosyl-L-methionine + vanillate
methyl vanillate + S-adenosyl-L-homocysteine
12% activity compared to salicylate
-
-
?
S-adenosyl-L-methionine + vanillic acid
S-adenosyl-L-homocysteine + methyl 4-hydroxy-3-methoxybenzoate
5.1% activity of wild-type enzyme compared to salicylate methylation
-
-
?
S-adenosyl-L-methionine + 4-hydroxybenzoate
S-adenosyl-L-homocysteine + methyl 4-hydroxybenzoate
-
lower catalytic efficiency with 4-hydroxybenzoate compared to salicylate
-
-
?
S-adenosyl-L-methionine + salicylate
S-adenosyl-L-homocysteine + methyl salicylate
-
-
-
-
?
S-adenosyl-L-methionine + 3-hydroxybenzoic acid
S-adenosyl-L-homocysteine + methyl 3-hydroxybenzoate
less than 2% relative activity at 1 mM methyl acceptor compared to activity with salicylate set at 100%
-
-
?
S-adenosyl-L-methionine + 3-hydroxybenzoic acid
S-adenosyl-L-homocysteine + methyl 3-hydroxybenzoate
17% activity of wild-type enzyme compared to salicylate methylation
-
-
?
S-adenosyl-L-methionine + salicylate
methyl salicylate + S-adenosyl-L-homocysteine
-
-
-
?
S-adenosyl-L-methionine + salicylate
methyl salicylate + S-adenosyl-L-homocysteine
best substrate
-
-
?
S-adenosyl-L-methionine + salicylate
S-adenosyl-L-homocysteine + methyl salicylate
-
-
-
?
S-adenosyl-L-methionine + salicylate
S-adenosyl-L-homocysteine + methyl salicylate
-
-
-
-
?
additional information
?
-
enzyme is also active with benzoic acid resulting in methyl benzoate formation
-
-
?
additional information
?
-
no activity with 4-hydroxybenzoic acid, vanillic acid, caffeic acid, p-coumaric acid and (+/-)jasmonic acid
-
-
?
additional information
?
-
no measurable methylation of jasmonic acid by wild-type SAMT using concentrations up to 5 mM
-
-
?
additional information
?
-
-
no measurable methylation of jasmonic acid by wild-type SAMT using concentrations up to 5 mM
-
-
?
additional information
?
-
wild-type is also able to methylate benzoic acid with 48% activity compared to salicylate methylation
-
-
?
additional information
?
-
-
wild-type is also able to methylate benzoic acid with 48% activity compared to salicylate methylation
-
-
?
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
-
-
-
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
S-adenosyl-L-methionine + salicylate
methyl salicylate + S-adenosyl-L-homocysteine
-
-
-
?
S-adenosyl-L-methionine + salicylate
S-adenosyl-L-homocysteine + methyl salicylate
-
-
-
?
S-adenosyl-L-methionine + 4-hydroxybenzoate
S-adenosyl-L-homocysteine + methyl 4-hydroxybenzoate
-
lower catalytic efficiency with 4-hydroxybenzoate compared to salicylate
-
-
?
S-adenosyl-L-methionine + salicylate
S-adenosyl-L-homocysteine + methyl salicylate
-
-
-
-
?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.009
S-adenosyl-L-methionine
app. Km-value of nontagged and His-tagged protein, pH 7.5, 20°C
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
7.5
pH RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
6 - 9
at pH 6.0 activity is 80% of optimal activity, at pH 8.0 it is 75%, and at pH 9 it is less than 50%
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SOURCE
highest SAMT activity compared to the levels of activity in all other floral parts or the vegetative tissue
UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
SEQUENCE
LOCALIZATION PREDICTION
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable).
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable). SAMT_CLABR
359
0
40289
Swiss-Prot
other Location (Reliability: 3)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
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
SAMT crystallized from ammonium sulfate solution, model spanned the entire 359 residues of full-length Clarkia SAMT
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Y147S
reduced methylation of benzoic acid and 3-hydroxybenzoic acid but slightly increased activity towards jasmonic acid
Y147S/M150H
significant increase in the ability to turn over jasmonic acid and vanillic acid
Y147S/M150H/F347Y
slightly increased activity towards short-chain carboxylic acids and jasmonic acid
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
Y147S/M150H/I225Q/F347Y
greatest specific activities against 3-hydroxybenzoic acid, vanillic acid and jasmonic acid but reduced activity towards salicylic acid
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
20 - 40
SAMT is 100% stable for 30 min at 20°C and 80% stable for 30 min at 30°C, but after 30 min incubation at 40°C it completely loses activity
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
by DE53 DEAE anion exchange, HAP and Mono-Q anion exchange chromatography
N-terminal polyhistidine-tagged SAMT protein purified by Ni2+ affinity chromatography and gel filtration chromatography using a Superdex-75 column
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
Clarkia breweri SAMT cloned into expression vector pET28a(+) and construct transformed into Escherichia coli BL21(DE3) cells
SAMT cDNA expressed with pET-11a/pET-28 vector in Escherichia coli which synthesizes a functional non-tagged/His-tagged SAMT protein
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Ross, J.R.; Nam, K.H.; D'Auria, J.C.; Pichersky, E.
S-Adenosyl-L-methionine:salicylic acid carboxyl methyltransferase, an enzyme involved in floral scent production and plant defense, represents a new class of plant methyltransferases
Arch. Biochem. Biophys.
367
9-16
1999
Clarkia breweri (Q9SPV4)
Zubieta, C.; Ross, J.R.; Koscheski, P.; Yang, Y.; Pichersky, E.; Noel, J.P.
Structural basis for substrate recognition in the salicylic acid carboxyl methyltransferase family
Plant Cell
15
1704-1716
2003
Clarkia breweri (Q9SPV4), Clarkia breweri
Yao, J.; Xu, Q.; Chen, F.; Guo, H.
QM/MM free energy simulations of salicylic acid methyltransferase: effects of stabilization of TS-like structures on substrate specificity
J. Phys. Chem. B
115
389-396
2011
Clarkia breweri
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