Study of the Role of Purine Phosphoribosyltransferases in the Uptake of Adenine and Guanine by Schizosaccharomyces pombe Cells

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Abstract
In the yeast S. pombe 972h- the uptake rate of adenine and guanine is related to variations in the specific activity of the corresponding phosphoribosyltransferases during the growth of the culture. The mutant strains dap1, devoid of adenine phosphoribosyltransferase activity, and pur1, devoid of guanine phosphoribosyltransferase activity have a lowered uptake rate of adenine and guanine, respectively, along with an increased apparent Km value for these purines in comparison to the wild-type 972h-. The uptake rate of the purines is strongly dependent on the pH of the uptake medium in 972h- as well as in the strains dap1 and pur1, the optimum being between pH 4 and pH 5. A new method of extraction of 5-phosphoribosyl-1-pyrophosphate [P-Rib-P2] from the yeast was devised. Important fluctuations of the P-Rib-P2 pool were measured in S. pombe at different stages of growth, the maximum taking place at the start of the exponential phase, whereas no variations in the specific activity of the P-Rib-P2 synthetase could be observed during the growth. The P-Rib-P2 intracellular content in the mutants devoid of purine phosphoribosyltransferases, namely pur1, dap1 and pur1, dap1, was increased up to 5-fold as compared to the wild-type strain. The effect of intracellular concentrations of P-Rib-P2, a substrate for phosphoribosyltransferases, on the uptake rate of purines was studied: addition of formycin to the growth medium lowered simultaneously the P-Rib-P2 intracellular content and the uptake of adenine and guanine. Although the results demonstrate the activating effect of phosphoribosyltransferase activities on the uptake of adenine and guanine, they do not support the hypothesis of a group translocation mechanism.