Characterization of cyanogen bromide peptides of hypoxanthine phosphoribosyltransferase by a micromethod

Human hypoxanthine phosphoribosyltransferase (HPRT-EC 2.4.2.8), either from erythrocytes labeled in vitro with 125I or from HeLa cells labeled in vivo with [3H]leucine, [3H]lysine, or [3H]arginine, was cleaved with cyanogen bromide. All four labeled enzymes produced cyanogen bromide peptides with molecular weights of 7800, 5600, 4100, 2500, and 1300 which were separated by sodium dodecyl sulfate urea polyacrylamide gel electrophoresis. Experiments with [35S]methionine-labeled HPRT indicated that 88 to 94% of the 35S label was released by the cyanogen bromide reaction. The relative distribution of radioactivity in each of the tritiated peptides indicates that there are 20 leucines, 15 lysines, and 12 arginines per enzyme subunit