ADENYLATE AND 2,6-DIAMINOPURINE RIBONUCLEOTIDE PYROPHOSPHORYLASE ACTIVITIES OF L CELLS

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Abstract
The reaction kinetics of adenylate (AMP) and 2,6-diaminopurine ribonucleoside 5′-monophosphate (DAPRP) pyrophosphorylase activities of extracts of L-strain mouse fibroblast cells were studied by chromatographic and radioactive tracer techniques. The apparent Kmvalues found for 5-phosphoribosyl-α-1-pyrophosphate (PRPP) were 6.9 × 10−5 M for AMP pyrophosphorylase activity and 3.6 × 10−4 M for DAPRP pyrophosphorylase activity. The apparent Kmfound for adenine was 3.0 × 10−5 M, whereas the apparent Kmcalculated for 2,6-diaminopurine was 1.8 × 10−3 M. An optimum pH of 7.6 was found for AMP pyrophosphorylase, whereas an optimum pH range of 7.6–7.9 was found for DAPRP pyrophosphorylase activity. Under assay conditions of zero-order kinetics, the AMP pyrophosphorylase activities of L-cell extracts were approximately twice the DAPRP pyrophosphorylase activities of the same extracts.AMP and DAPRP were found to be the major products of the AMP and DAPRP pyrophosphorylase reactions, respectively. DAPRP was analyzed chemically and spectrophotometrically, and absorption spectra at different pH values are presented.The effects of sulfate ions, storage, and enzyme inactivation techniques on enzyme activities were also studied.