The Uptake of Adenine by Isolated Human Platelet Membranes

Summary

Adenine uptake into isolated platelet membranes had about the same Km (151 ± 21 • 9 nM) as uptake into intact cells (159 ± 21 nM) and was also competitively inhibited by papaverine and hypoxanthine. No uptake occurred at 0° and accumulated adenine was converted to AMP. AMP was not firmly bound to protein as judged by chromatography of triton X-100 solubilized membranes on Sephadex G25. The pH optimum for adenine uptake was at pH 5-5. Exogenous 5-phosphoribosyl-l-pyrophos- phate strongly stimulated uptake. These data may be explained by uptake of adenine by facilitated diffusion followed by conversion to AMP by adenine phosphoribosyltransferase but group translocation cannot be entirely excluded.

^* On leave of absence from the Division of Haemostasis, Dept, of Intern. Med., State University Hosp., Utrecht, The Netherlands.

• References

• 1 Day H. J, Holmsen H, Hovig T. 1969; Subcellular particles of human platelets. Scandinavian Journal of Haematology, supplementum 07: 3.
• 2 French P. C, Holmsen H, Stormorken H. 1970; Adenine nucleotide metabolism of blood platelets. VII. ATPases: subcellular localization and behaviour during the thrombin-platelet interaction. Biochimica Biophysica Acta 206: 438.
• 3 French P. C, Holme R. 1974; A method for blood platelet homogenization using the Aminco-French pressure cell. Thrombosis et Diathesis Hæmorrhagica 32: 432.
• 4 Gaintner J. R, Jackson D. P, Maynert E. W. 1962; The action of thrombin on platelet 5-hydroxytryptamine. Bulletin of the Johns Hopkins Hospital 111: 185.
• 5 Hochstadt-Ozer J, Stadtman E. R. 1971; The regulation of purine utilization in bacteria. I. Purification of adenine phosphoribosyltransferase from E. coli K₁₂ and control of activity of nucleotides. Journal of Biological Chemistry 246: 5294.
• 6 Hochstadt-Ozer J, Stadtman E. R. 1971; The regulation of purine utilization in bacteria. II. Adenine phosphoribosyl-transferase in isolated membrane preparations and its role in transport of adenine across the membrane. Journal of Biological Chemistry 246: 5304.
• 7 Hochstadt-Ozer J, Stadtman E. R. 1971; The regulation of purine utilization in bacteria. III. The involvement of purine phosphoribosyltransferases in the uptake of adenine and other nucleic acid precursors by intact resting cells. Journal of Biological Chemistry 246: 5312.
• 8 Holmsen H, Rozenberg M. C. 1968; Adenine nucleotide metabolism of blood platelets. III. Adenine phosphoribosyl transferase and nucleotide formation from exogenous adenine. Biochimica Biophysica Acta 157: 266.
• 9 Lage B. A, Scrutton M. C. 1973 personal communication.
• 10 Miller G. L. 1959; Protein determinations for a large number of samples. Analytical Chemistry 31: 964.
• 11 Schacter H. 1973; On the interpretation of Michælis constants for transport. Journal of Biological Chemistry 248: 974.
• 12 Sixma J. J, Holmsen H, andTrieschnigg A. C. M. 1973; Adenine nucleotide metabolism of blood platelets. VIII. Transport of adenine into human platelets. Biochimica Biophysica Acta 298: 460.