| Size | Price | |
|---|---|---|
| 500mg | ||
| 1g | ||
| Other Sizes |
| Targets |
The direct target of (E)-2-Hexenoyl-CoA in this study is the enzyme RevT (a crotonyl-CoA carboxylase/reductase homolog). Kinetic parameters for RevT using (E)-2-Hexenoyl-CoA as a substrate are reported as: Michaelis constant
Km
=
0.27
±
0.01
K
m
=0.27±0.01 mM, turnover number
Kcat
=
28.3
±
0.5
k
cat
=28.3±0.5 min
−
1
−1
, and catalytic efficiency
Kcat/Km
=
104
k
cat
/K
m
=104 mM
−
1
−1
min
−
1
−1
. NADPH is a co-substrate for this reaction, with a
Km
of
0.12
±
0.01
0.12±0.01 mM and
Kcat/Km
=
282
k
cat
/K
m
=282 mM
−
1
−1
min
−
1
−1
when using (E)-2-hexenoyl-CoA. [1]
|
|---|---|
| ln Vitro |
The in vitro activity of (E)-2-Hexenoyl-CoA is defined by its enzymatic conversion by RevT. In a spectrophotometric assay, RevT catalyzes the carboxylation and reduction of (E)-2-hexenoyl-CoA in the presence of NADPH and NaHCO
3
3
, producing butylmalonyl-CoA. The initial rate of the reaction is measured by monitoring the oxidation of NADPH at 340 nm. Furthermore, in a coupled enzyme assay containing RevS and RevT, (E)-2-hexenoic acid is first converted to (E)-2-hexenoyl-CoA, which is then directly transformed into butylmalonyl-CoA, demonstrating the complete in vitro biosynthesis of this extender unit from the free fatty acid. [1]
|
| Enzyme Assay |
To synthesize (E)-2-Hexenoyl-CoA for biochemical assays, a transesterification method was employed. First, thiophenyl (E)-2-hexenoate was synthesized by reacting (E)-2-hexenoic acid with thiophenol using dicyclohexylcarbodiimide as a condensing agent. For the enzymatic assay or preparation, a solution of this thiophenyl ester in tetrahydrofuran was added to a stirred solution of free CoA in 0.1 M NaHCO
3
3
at 0°C. The mixture was allowed to stir overnight at 4°C. The product was then purified using Cosmosil column chromatography with a gradient of water/methanol, followed by lyophilization to obtain an amorphous powder. [1]
The kinetic parameters of RevT for (E)-2-Hexenoyl-CoA were determined using a spectrophotometric NADPH oxidation assay. The reaction mixture (200 µL) contained 100 mM Tris-HCl (pH 7.5), 10% glycerol, 10 mM NaHCO
3
3
, 11 mM MgCl
2
2
, 1 mM EDTA, varying concentrations of (E)-2-hexenoyl-CoA (0.1 to 2 mM), 0.3 mM NADPH, and 1 µM RevT. The reaction was initiated by adding RevT after a 3-minute pre-incubation at 25°C. The decrease in absorbance at 340 nm, corresponding to NADPH oxidation, was measured. The kinetic constants were calculated via nonlinear regression fit to the Michaelis-Menten equation. [1]
|
| References | |
| Additional Infomation |
(E)-2-Hexenoyl-CoA is a physiological substrate for the crotonyl-CoA carboxylase/reductase (CCR) family of enzymes. In this study, it was used to demonstrate that RevT, a CCR homolog, functions as a carboxylase/reductase on CoA-linked substrates. A feeding experiment using [1-
13
13
C]hexanoic acid was performed on the ΔrevR mutant of Streptomyces sp. SN-593. The subsequent
13
13
C NMR analysis of the isolated reveromycin A showed a highly enriched signal at 25.4 ppm (assigned to C17), confirming that exogenously supplied hexanoic acid – which can be activated to (E)-2-hexenoyl-CoA – is directly utilized for the biosynthesis of the butylmalonyl-CoA extender unit and incorporated into the final polyketide structure. [1]
|
| Molecular Formula |
C27H44N7O17P3S
|
|---|---|
| Molecular Weight |
863.66
|
| Exact Mass |
863.173
|
| CAS # |
10018-93-6
|
| PubChem CID |
440612
|
| Appearance |
Typically exists as solid at room temperature
|
| LogP |
1.384
|
| Hydrogen Bond Donor Count |
9
|
| Hydrogen Bond Acceptor Count |
22
|
| Rotatable Bond Count |
23
|
| Heavy Atom Count |
55
|
| Complexity |
1490
|
| Defined Atom Stereocenter Count |
4
|
| SMILES |
CCCC=CC(=O)SCCNC(=O)CCNC(=O)C(C(C)(C)COP(=O)(O)OP(=O)(O)OC[C@@H]1[C@H]([C@H]([C@@H](O1)N2C=NC3=C(N=CN=C32)N)O)OP(=O)(O)O)O
|
| InChi Key |
OINXHIBNZUUIMR-ZOGSZLKASA-N
|
| InChi Code |
InChI=1S/C27H44N7O17P3S/c1-4-5-6-7-18(36)55-11-10-29-17(35)8-9-30-25(39)22(38)27(2,3)13-48-54(45,46)51-53(43,44)47-12-16-21(50-52(40,41)42)20(37)26(49-16)34-15-33-19-23(28)31-14-32-24(19)34/h6-7,14-16,20-22,26,37-38H,4-5,8-13H2,1-3H3,(H,29,35)(H,30,39)(H,43,44)(H,45,46)(H2,28,31,32)(H2,40,41,42)/t16-,20-,21-,22?,26-/m1/s1
|
| Chemical Name |
S-[2-[3-[[4-[[[(2R,3S,4R,5R)-5-(6-aminopurin-9-yl)-4-hydroxy-3-phosphonooxyoxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-hydroxyphosphoryl]oxy-2-hydroxy-3,3-dimethylbutanoyl]amino]propanoylamino]ethyl] hex-2-enethioate
|
| Synonyms |
(E)-2-Hexenoyl-CoA;
trans-Hex-2-enoyl-CoA; 2E-hexenoyl-CoA; 2E-hexenoyl-coenzyme A; trans-2-Hexenoyl-CoA;
|
| HS Tariff Code |
2934.99.9001
|
| Storage |
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month |
| Shipping Condition |
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
|
| Solubility (In Vitro) |
Typically soluble in DMSO (e.g. 10 mM)
|
|---|---|
| Solubility (In Vivo) |
Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.
Injection Formulations
Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)(e.g. IP/IV/IM/SC) *Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution. Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline) Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil) Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals). View More
Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)] Oral Formulations
Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium) Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals). View More
Oral Formulation 3: Dissolved in PEG400 (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 1.1579 mL | 5.7893 mL | 11.5786 mL | |
| 5 mM | 0.2316 mL | 1.1579 mL | 2.3157 mL | |
| 10 mM | 0.1158 mL | 0.5789 mL | 1.1579 mL |
*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.
Calculation results
Working concentration： mg/mL；
Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution concentration mg/mL). Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug.
Method for preparing in vivo formulation:：Take μL DMSO stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O，mix and clarify.
(1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.
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