AMIDOPHOSPHORIBOSYLTRANSFERASE LIMITS THE RATE OF CELL GROWTH-LINKED DE-NOVO PURINE BIOSYNTHESIS IN THE PRESENCE OF CONSTANT CAPACITY OF SALVAGE PURINE BIOSYNTHESIS

Factors controlling relative flux rates of the de novo and salvage pat hways of purine nucleotide biosynthesis during animal cell growth are not fully understood, To examine the relative role of each pathway for cell. growth, three cell lines including CHO K1 (a wild-type Chinese hamster ovary fibroblast cell line), CHO ade (-)A (an auxotrophic cell line deficient of amidophosphoribosyltransferase (ATase), a presumed rate-limiting enzyme of the de novo pathway), and CHO ade (-)A transfe cted with human ATase cDNA ((-)A+hATase) resulting in 30-350% of the A Tase activity of CHO K1, were cultured in purine-rich or purine-free m edia, Based on the enzyme activities of ATase and hypoxanthine phospho ribosyltransferase, the metabolic rate of the de novo and salvage path ways, the rate of cell growth (growth rate) in three cell lines under various culture conditions, and the effect of hypoxanthine infusion on the metabolic rate of the de novo pathway in rat liver, we concluded the following, 1) In (-)A+hATase transfectants, ATase activity limits the rate of the de novo pathway, which is closely linked with the grow th rate, 2) Purine nucleotides are synthesized preferentially by the s alvage path way as long as hypoxanthine, the most essential source of purine salvage, can be utilized, which was confirmed in rat liver in v ivo by hypoxanthine infusion, The preferential usage of the salvage pa thway results in sparing the energy expenditure required for de novo s ynthesis, 3) The regulatory capacity of the de novo pathway (about 200 %) was larger than that of the salvage pathway (about 20%) with consta nt hypoxanthine phosphoribosyltransferase activity.