{"product_id":"phosphorylase-b-enzyme-activity-tbp0084","title":"Phosphorylase-b, 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\u003c\/p\u003e\n\u003cp\u003e\u003cu\u003eSolubility:\u003c\/u\u003e Soluble in distilled water or dilute buffer\u003c\/p\u003e\n\u003cp\u003e\u003cu\u003eStability:\u003c\/u\u003e -20° C; -4° F\u003c\/p\u003e\n\u003cp\u003e\u003cu\u003eActivity:\u003c\/u\u003e 60 U\/mg protein\u003c\/p\u003e\n\u003cp\u003e\u003cu\u003eProtein:\u003c\/u\u003e 90%\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\u003eThat amount of enzyme which will liberate one micromole of glucose-1-phosphate from glycogen and orthophosphate, in the presence of 5′-AMP, per minute at pH 6.8 at 37°C.\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\u003eThe assay method which is described.\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\u003ePhosphorylases (EC 2.4.1.1.) catalyze the breakdown of glycogen in liver and muscle. Similar enzymes, occurring in plants, catalyze the hydrolysis of starch. The hydrolysis of glycogen is represented by the following reaction:\u003c\/p\u003e\n\u003cp\u003e \u003c\/p\u003e\n\u003cp\u003eThe phosphorylase of skeletal muscle occurs in two forms, the active form (phosphorylase-a) and a much less active form (phosphorylase-b). These two forms are interconvertible. Rabbit muscle phosphorylase has been extensively studied. Phosphorylase-a has a molecular weight of 370,000 and phosphorylase-b has a molecular weight of 185,000.\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\u003e0.05 M Potassium phosphate buffer, pH 6.8 containing 0.1 mM EDTA.\u003c\/li\u003e\n\u003cli\u003e0.1 M Magnesium chloride in distilled water.\u003c\/li\u003e\n\u003cli\u003eNADP (10 mg\/ml) in distilled water. Prepare fresh.\u003c\/li\u003e\n\u003cli\u003eGlycogen (6 mg\/ml) in buffer. Prepare fresh.\u003c\/li\u003e\n\u003cli\u003e1 mM adenosine 5′-monophosphate (sodium salt) in buffer. Prepare fresh.\u003c\/li\u003e\n\u003cli\u003ePhosphoglucomutase (PGM) solution. Prepare a solution in cold 1% bovine serum albumin to yield a concentration of 20 U\/ml. Must be prepared fresh.\u003c\/li\u003e\n\u003cli\u003eGlucose-6-P-dehydrogenase (G6P-DH) solution. Prepare a solution in cold 1% bovine serum albumin to yield a final concentration of 100 U\/ml. Must be prepared fresh.\u003c\/li\u003e\n\u003cli\u003ePhosphorylase (enzyme) solution. Prepare a solution in cold 1% bovine serum albumin to yield a final concentration of 2-4 U\/ml. Must be prepared fresh immediately prior to assay. (Note: the enzyme preparation contains 5′-AMP which is needed for its catalytic activity.)\u003c\/li\u003e\n\u003c\/ol\u003e\n\u003cp\u003e \u003c\/p\u003e\n\u003cp\u003e\u003cspan style=\"text-decoration: underline\"\u003e\u003cstrong\u003e\u003cspan style=\"font-size: 18px\"\u003eProcedure\u003c\/span\u003e\u003c\/strong\u003e\u003c\/span\u003e\u003c\/p\u003e\n\u003col\u003e\n\u003cli\u003eSet spectrophotometer (equipped with strip chart recorder and temperature control) at 340 nm and 37°C.\u003c\/li\u003e\n\u003cli\u003eInto a cuvette pipette the following reagents:\u003c\/li\u003e\n\u003c\/ol\u003e\n\u003cp style=\"padding-left: 40px\"\u003ePotassium phosphate buffer with EDTA 2.4 ml\u003c\/p\u003e\n\u003cp style=\"padding-left: 40px\"\u003eMagnesium chloride 0.1 ml\u003c\/p\u003e\n\u003cp style=\"padding-left: 40px\"\u003eNADP 0.1 ml\u003c\/p\u003e\n\u003cp style=\"padding-left: 40px\"\u003eGlycogen 0.1 ml\u003c\/p\u003e\n\u003cp style=\"padding-left: 40px\"\u003eAdenosine 5′-monophosphate 0.1 ml\u003c\/p\u003e\n\u003cp style=\"padding-left: 40px\"\u003ePhosphoglucomutase 0.1 ml\u003c\/p\u003e\n\u003cp style=\"padding-left: 40px\"\u003eGlucose-6-P-dehydrogenase 0.1 ml\u003c\/p\u003e\n\u003cp style=\"padding-left: 40px\"\u003eMix and incubate at 37¡C for 5 minutes in the spectrophotometer to attain temperature equilibration.\u003c\/p\u003e\n\u003col start=\"3\"\u003e\n\u003cli\u003eEstablish blank rate, if any, at 340 nm.\u003c\/li\u003e\n\u003cli\u003eInitiate the reaction by adding 0.1 ml phosphorylase (enzyme) solution. Record the change in absorbance at 340 nm for 5-8 minutes.\u003c\/li\u003e\n\u003cli\u003eCalculate µE340\/min\u003c\/li\u003e\n\u003c\/ol\u003e\n\u003cp\u003e\u003c!--TBP0084-10MG TBP0084-50MG--\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\/TBP0084-Phosphorylase-b-1.pdf\"\u003eDatasheet (PDF)\u003c\/a\u003e\u003c\/li\u003e\n\u003cli\u003e\u003ca href=\"https:\/\/tribioscience.com\/wp-content\/uploads\/2024\/03\/TBP0084_Phosphorylase-b-Safety-Data-Sheet-1.pdf\"\u003eMSDS (PDF)\u003c\/a\u003e\u003c\/li\u003e\n\u003c\/ul\u003e","brand":"TriBioScience","offers":[{"title":"10 mg","offer_id":45986500477007,"sku":"TBP0084-10MG","price":0.0,"currency_code":"ILS","in_stock":true},{"title":"50 mg","offer_id":45986500509775,"sku":"TBP0084-50MG","price":0.0,"currency_code":"ILS","in_stock":true}],"thumbnail_url":"\/\/cdn.shopify.com\/s\/files\/1\/0712\/0748\/5519\/files\/TBP0084-1.png?v=1782812676","url":"https:\/\/www.danyel.co.il\/products\/phosphorylase-b-enzyme-activity-tbp0084","provider":"danyel biotech","version":"1.0","type":"link"}