Purification and Characterization of the Hypoxanthine‐Guanine Phosphoribosyltransferase from Saccharomyces cerevisiae

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Abstract
Hypoxanthine-guanine phosphoribosyltransferase (EC 2.4.2.8) from S. cerevisiae was purified 9400-fold by affinity chromatography giving rise to an electrophoretically homogeneous preparation. The MW of the enzyme was determined by gel filtration with Sephadex G-100 and by sodium dodecylsulfate gel electrophoresis. Both methods reveal a MW of 51,000. The enzyme requires Mg2+ and has its pH optimum at 8.5. Isoelectric focusing and gel electrophoresis of the purified extract reveals a single band which exhibits enzyme activity. The isoelectric point of the enzyme is 5.1. The enzyme displays Michaelis-Menten kinetics with apparent Km for hypoxanthine, guanine and phosphoribosylpyrophosphate of 23 .mu.M, 18 .mu.M and 50 .mu.M, respectively.