Purine biosynthesis de novo in bovine retina: purification and characterization of amidophosphoribosyl transferase and phosphoribosyl pyrophosphate synthetase.
The ability of bovine retina to synthesize purines de novo is shown for the first time. Amidophosphoribosyl transferase (EC 2.4.2.14), the enzyme controlling the rate of the process, and phosphoribosyl pyrophosphate synthetase (EC 2.7.6.1), the enzyme regulating the intracellular contents of phosphoribosyl pyrophosphate (PRPP), were purified and characterized. The molecular masses of the enzyme subunits are similar to those of the purified enzyme from the liver. The molecular masses of amidophosphoribosyl transferase, PRPP synthetase catalytic subunit, and two PRPP synthetase-associated proteins are 50, 34, 39, and 41 kD, respectively. The apparent Km values of the enzymes and coenzymes are similar to those of the purified enzymes from the liver. For amidophosphoribosyl transferase, the apparent Km for Gln and PRPP are 0.75 +/- 0.05 and 0.66 +/- 0.09 mM, respectively (the corresponding Vmax values are 59 +/- 3 and 136 +/- 12 nmoles PPi/min per mg protein). For PRPP synthetase, the apparent Km for ribose-5-phosphate and ATP are 37.9 +/- 0.5 and 53 +/- 7 microM, respectively (the corresponding Vmax values are 61 +/- 4 and 52 +/- 3 nmoles PRPP/min per mg protein). The sensitivity of the retinal PRPP synthetase to inhibition by ADP and AMP was significantly lower than that of the enzyme from the liver.
Sechenov Institute of Evolutionary Physiology and Biochemistry, Russian Academy of Sciences, St. Petersburg, 194223, Russia. julia@snk1.mail.iephb.ru
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D000582:Amidophosphoribosyltransferase / Q000737:chemistry* / Q000302:isolation & purification*; D000818:Animals; D002417:Cattle; D002847:Chromatography, Agarose; D002848:Chromatography, DEAE-Cellulose; D007700:Kinetics; D011687:Purines / Q000096:biosynthesis*; D012160:Retina / Q000378:metabolism*; D012268:Ribose-Phosphate Pyrophosphokinase / Q000737:chemistry* / Q000302:isolation & purification*
D011687:Purines; D000582:Amidophosphoribosyltransferase; D012268:Ribose-Phosphate Pyrophosphokinase