Abstract
An unusual new purine-requiring mutant, Ade −P AB, of Chinese hamster ovary cells (CHO-K1) is described. Ade −P AB will grow in medium supplemented with hypoxanthine, adenine, or aminoimidazole carboxamide. Ade −P AB fails to show genetic complementation with either Ade −A, defective in amidophosphoribosyltransferase (E.C. 2.4.2.14), or Ade −B, defective in phosphoribosylformylglycinamidine FGAM) synthetase (E.C. 6.3.5.3.), but will complement all five of our other hypoxanthine-requiring Ade − complementation groups. Analysis of purine synthesis in wild-type, mutant, and revertant cells and analysis of relevant enzyme activities in cell-free extracts prepared from these cells demonstrates that Ade −P AB is similar to Ade −B in that it has lost FGAM synthetase activity, and is similar to Ade −A in that it has lost glutamine-dependent amidophosphoribosyltransferase activity. Unlike Ade −A, however, Ade −P AB retains the ability to synthesize phosphoribosylamine (PRA), the product of the amidophosphoribosyltransferase reaction, if NH 4Cl is substituted for glutamine as the nitrogen donor. Moreover, partial revertants of Ade −P AB can apparently synthesize sufficient purines for growth using the NH 4Cl-dependent reaction. The available evidence indicates that neither a double mutation nor a deletion is probable in Ade −P AB. We discuss the relevance of these observations for our understanding of both the regulation of purine biosynthesis in mammalian cells and the structural organization of the enzymes defective in Ade −P AB and the genes coding for these enzymes.
Published Version
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