Purine nucleoside phosphorylase from human erythrocytes: physicochemical properties of the crystalline enzyme

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Abstract
The major physicochemical properties of human erythrocytic purine nucleoside phosphorylase (PNPase) are described. The MW estimated by ultracentrifugation, molecular sieving and sucrose density gradient centrifugation, ranged from 87,000-92,000. Other physical constants of erythrocytic PNPase were: sedimentation coefficient (S20,W), 5.4 S obtained by sedimentation analysis and 5.5 S by the sucrose density gradient procedure; Stokes radius, 38 .ANG.; calculated diffusion coefficient (D20,W), 5.7 .times. 10-7 cm2 s-1; frictional ratio, 1.29; and partial specific volume calculated from amino acid analysis, 0.73 cm3 g-1. The circular dichroism spectra of the human erythrocytic and bovine spleen PNPase were almost identical and indicated a very low .alpha.-helical content. Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate indicated that the MW of the PNPase subunit is 30,000 .+-. 500. These results corroborate earlier reports that the native enzyme is a homologous trimer. Comparative studies with crystalline bovine spleen PNPase confirmed that it is also a trimer but is somewhat smaller than the human erythrocytic enzyme with a MW of about 86,000.