{"product_id":"catechol-o-methyl-transferase-enzyme-activity-tbp0061","title":"Catechol O-Methyl Transferase, Enzyme Activity","description":"\u003cp\u003e\u003cspan style=\"font-size: 18px\"\u003e\u003cstrong\u003e\u003cu\u003eProduct Details\u003c\/u\u003e\u003c\/strong\u003e\u003c\/span\u003e\u003c\/p\u003e\n\u003cp\u003e\u003cu\u003eForm:\u003c\/u\u003e Freeze-dried powder\u003c\/p\u003e\n\u003cp\u003e\u003cu\u003eSolubility:\u003c\/u\u003e Distilled water or dilute buffer\u003c\/p\u003e\n\u003cp\u003e\u003cu\u003eStability:\u003c\/u\u003e Store at -20° C (-4° F)\u003c\/p\u003e\n\u003cp\u003e\u003cu\u003eActivity:\u003c\/u\u003e 4000 U\/mg protein\u003c\/p\u003e\n\u003cp\u003e\u003cu\u003eProtein:\u003c\/u\u003e 90%\u003c\/p\u003e\n\u003cp\u003e\u003cu\u003eContaminants:\u003c\/u\u003e PNMT \u0026lt;0.01%\u003c\/p\u003e\n\u003cp\u003e \u003c\/p\u003e\n\u003cp\u003e\u003cspan style=\"font-size: 18px\"\u003e\u003cstrong\u003e\u003cu\u003eUnit Definition\u003c\/u\u003e\u003c\/strong\u003e\u003c\/span\u003e\u003c\/p\u003e\n\u003cp\u003eThe amount of enzyme which will catalyze the methylation of one nanomole of dihydroxybenzoic acid per hour at 37°C, pH 8.0, using S-adenosyl-L-(methyl 14C)-methionine as the methyl donor.\u003c\/p\u003e\n\u003cp\u003e \u003c\/p\u003e\n\u003cp\u003e\u003cspan style=\"font-size: 18px\"\u003e\u003cstrong\u003e\u003cu\u003eAssay Method\u003c\/u\u003e\u003c\/strong\u003e\u003c\/span\u003e\u003c\/p\u003e\n\u003cp\u003eEnzyme activity is determined by measuring the amount of 14C incorporated into the methylated substrate (3,4- Dihydroxybenzoic acid).\u003c\/p\u003e\n\u003cp\u003e \u003c\/p\u003e\n\u003cp\u003e\u003cspan style=\"font-size: 18px\"\u003e\u003cstrong\u003e\u003cu\u003eApplications\u003c\/u\u003e\u003c\/strong\u003e\u003c\/span\u003e\u003c\/p\u003e\n\u003cp\u003eThe enzyme, catechol O-methyltransferase (COMT) (EC 2.1.1.6) can O-methylate a wide variety of substituted catechols such as dopamine, epinephrine and norepinephine. This is an important step in the metabolic detoxification of catecholamines. A representative reaction is shown below:\u003c\/p\u003e\n\u003cp\u003e \u003c\/p\u003e\n\u003cp\u003e\u003cspan style=\"font-size: 18px\"\u003e\u003cstrong\u003e\u003cu\u003eReagents\u003c\/u\u003e\u003c\/strong\u003e\u003c\/span\u003e\u003c\/p\u003e\n\u003col\u003e\n\u003cli\u003e1 mM S-Adenosylmethionine (SAM), (0.435 mg\/ml) 100µl. (This product must be of highest purity and must not contain traces of S-adenosylhomocysteine).\u003c\/li\u003e\n\u003cli\u003e10 mM Magnesium chloride, in distilled water (0.952 mg\/ml). 100µl.\u003c\/li\u003e\n\u003cli\u003e5 mM Dithiothreitol (0.61 mg\/ml) in distilled water. 100 µl.\u003c\/li\u003e\n\u003cli\u003e0.5 M Tris-HCl buffer, pH 8.0 in distilled water. 200 µl.\u003c\/li\u003e\n\u003cli\u003e10-50 mM 3,4 Dihydroxybenzoic acid, (1.56-7.81 mg\/ml) in water. 200 µl.\u003c\/li\u003e\n\u003cli\u003e14C S-Adenosylmethionine, (55mCi\/mMole). Distilled water is added to this solution to make 1 ml. It must be kept on ice until needed. Use 10µCi in the assay mixture.\u003c\/li\u003e\n\u003c\/ol\u003e\n\u003cp style=\"padding-left: 40px\"\u003eNote: The assay solution used for COMT assay is prepared by mixing the above reagents in the amounts indicated.\u003c\/p\u003e\n\u003col start=\"7\"\u003e\n\u003cli\u003e1% Bovine serum albumin (BSA) solution. Dissolve 1.0 g BSA in 100 ml distilled water. Albumin should be of highest purity.\u003c\/li\u003e\n\u003cli\u003eCOMT (enzyme) solution: Prepare a suitable dilution of the enzyme using cold 1% BSA. Prepare fresh prior to assay.\u003c\/li\u003e\n\u003cli\u003eHydrochloric acid (conc.).\u003c\/li\u003e\n\u003cli\u003eToluene: 3-methyl butanol, 7:3 (v\/v).\u003c\/li\u003e\n\u003c\/ol\u003e\n\u003cp\u003e \u003c\/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cspan style=\"text-decoration: underline\"\u003e\u003cspan style=\"font-size: 18px\"\u003eProcedure\u003c\/span\u003e\u003c\/span\u003e\u003c\/strong\u003e\u003c\/p\u003e\n\u003col\u003e\n\u003cli\u003ePipette 100 µl of the assay solution in the assay tube.\u003c\/li\u003e\n\u003cli\u003eAdd 100 µl of a suitable dilution of COMT (enzyme) to the assay tube at zero time. Mix.\u003c\/li\u003e\n\u003cli\u003eIncubate the assay tube for 10 min at 37°C.\u003c\/li\u003e\n\u003cli\u003eTerminate the reaction by adding 0.1 ml HCl to the assay tube.\u003c\/li\u003e\n\u003cli\u003eExtract radioactive products into 6 ml of a mixture of toluene: 3 methylbutanol (7:3 v\/v) by shaking for 20 seconds.\u003c\/li\u003e\n\u003cli\u003eThe aqueous phase is allowed to settle and then discarded, while the upper organic phase is saved and dried and then taken up in 1 ml ethanol and 14C radioactivity is counted in a scintillation spectrometer.\u003c\/li\u003e\n\u003c\/ol\u003e\n\u003cp\u003e\u003c!--TBP0061-1MG TBP0061-5MG--\u003e\u003c\/p\u003e\n\u003cp\u003e\u003cspan style=\"font-size: 13px; font-weight: 700; text-transform: uppercase;\"\u003e\u003cstrong\u003eResearch use only.\u003c\/strong\u003e\u003c\/span\u003e\u003c\/p\u003e\n\u003ch2\u003eDocuments\u003c\/h2\u003e\n\u003cul\u003e\n\u003cli\u003e\u003ca href=\"https:\/\/tribioscience.com\/wp-content\/uploads\/2024\/03\/TBP0061-Catechol-O-Methyl-Transferase.pdf\"\u003eDatasheet (PDF)\u003c\/a\u003e\u003c\/li\u003e\n\u003cli\u003e\u003ca href=\"https:\/\/tribioscience.com\/wp-content\/uploads\/2024\/03\/TBP0061_Catechol-O-Methyl-Transferase-Safety-Data-Sheet-1.pdf\"\u003eMSDS (PDF)\u003c\/a\u003e\u003c\/li\u003e\n\u003c\/ul\u003e","brand":"TriBioScience","offers":[{"title":"1 mg","offer_id":45986498281551,"sku":"TBP0061-1MG","price":0.0,"currency_code":"ILS","in_stock":true},{"title":"5 mg","offer_id":45986498314319,"sku":"TBP0061-5MG","price":0.0,"currency_code":"ILS","in_stock":true}],"thumbnail_url":"\/\/cdn.shopify.com\/s\/files\/1\/0712\/0748\/5519\/files\/TBP0061.png?v=1782812630","url":"https:\/\/www.danyel.co.il\/products\/catechol-o-methyl-transferase-enzyme-activity-tbp0061","provider":"danyel biotech","version":"1.0","type":"link"}