Membrane-associated purine metabolizing enzyme activities of human peripheral blood cells.
Sealed and unsealed plasma membrane vesicles were prepared from human erythrocytes and lymphocytes. Phosphoribosylpyrophosphate synthetase (PRibPP synthetase), hypoxanthine phosphoribosyltransferase (HPRTase), and adenine phosphoribosyltransferase (APRTase) activities are detectable on both inside-out and right-side-out sealed vesicles. Ghost preparations were about 0.2%, 1%, and 1.2% of the total erythrocyte and 0.5%, 5.3%, and 9.7% of the lymphocyte APRTase, HPRTase, and PRibPP synthetase activities. The rapid decrease in these enzyme activities, upon further purification of the membranes, seemed to suggest that they might be loosely bound extrinsic proteins. Evidence confirming the localization of these enzymes on the cell surface was obtained by measuring production of [14C]AMP by intact cells in medium containing [14C]adenine, ribose 5-phosphate, and Mg2+ATP. The formation of AMP was linear with time and number of cells present. Magnesium and phosphate exerted different effects on the production of extracellular AMP than on intracellular, which involves transport as well as phosphoribosylation. Cytosoluble and membrane-bound APRTase and PRibPP synthetase exhibited different catalytic properties and sensitivities to effectors. Membranes of erythrocytes of HPRTase-deficient patients contain little or no HPRTase activity when assayed in the absence of Triton. Reisolation of these membranes from admixture with normal hemolysates did not result in any bound activity; thus, the membrane-bound activity is not an artifact of the isolation procedure. Lysis with Triton released activity equal to about half that of control membranes. This is further evidence that the enzyme is firmly bound to the membrane.
D000228:Adenine Phosphoribosyltransferase / Q000097:blood; D002462:Cell Membrane / Q000201:enzymology; D004910:Erythrocyte Membrane / Q000201:enzymology*; D004912:Erythrocytes / Q000201:enzymology*; D006801:Humans; D007041:Hypoxanthine Phosphoribosyltransferase / Q000097:blood; D008214:Lymphocytes / Q000201:enzymology*; D008274:Magnesium / Q000494:pharmacology; D010710:Phosphates / Q000494:pharmacology; D011687:Purines / Q000097:blood*; D012268:Ribose-Phosphate Pyrophosphokinase / Q000097:blood; D013499:Surface Properties
D010710:Phosphates; D011687:Purines; D000228:Adenine Phosphoribosyltransferase; D007041:Hypoxanthine Phosphoribosyltransferase; D012268:Ribose-Phosphate Pyrophosphokinase; D008274:Magnesium