Evidence for genetically independent allosteric regulatory domains of the protein M1 subunit of mouse ribonucleotide reductase.
Ribonucleotide reductase is responsible for the reduction of the 2'-hydroxy moiety of all four ribonucleoside diphosphates to the corresponding deoxyribonucleotides. The overall activity of the enzyme is regulated by the allosteric effectors ATP (activator) and dATP (inhibitor), and the enzyme's substrate specificity is also controlled by nucleotide effectors. For instance, wild type ribonucleotide reductase from mouse T-lymphoma (S49) cells requires dGTP as a positive effector for ADP reduction. This effect of dGTP causes a reciprocal inhibition of CDP reduction. The dGuo-L mutant cell line, resistant to growth inhibition by exogenous deoxyguanosine, contains a nucleotide-binding subunit, protein M1, that conveys to its CDP reductase an insensitivity to dGTP (and dTTP) inhibition. The dGuo-L protein M1 also shows a decreased capacity to use ADP as a substrate, and therefore, the regulation of the substrate specificity is altered in the mutant protein M1. Another mutant cell line, dGuo-200-1, is resistant to deoxyadenosine and its ribonucleotide reductase is abnormally resistant to inhibition by dATP. The isolated mutant protein M1 from dGuo-200-1 cells has a CDP reductase activity which is stimulated by dATP, unlike the wild type enzyme which is inhibited by dATP. It appears that this mutant enzyme has lost the capacity to distinguish between dATP and ATP, but is still sensitive to regulation by dGTP and dTTP. Thus, the site of protein M1 regulating overall activity is altered in the dGuo-200-1 mutant, while the site regulating substrate specificity is normal. These characteristics of the mutants provide genetic evidence for two independent allosteric domains of protein M1, each responsible for a different aspect of nucleotide sensitivity of ribonucleotide reductase.
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000494:Allosteric Regulation; D000818:Animals; D002460:Cell Line; D003838:Deoxyadenine Nucleotides / Q000494:pharmacology; D007700:Kinetics; D008223:Lymphoma / Q000201:enzymology*; D046911:Macromolecular Substances; D051379:Mice; D009374:Neoplasms, Experimental / Q000201:enzymology; D012264:Ribonucleotide Reductases / Q000235:genetics* / Q000378:metabolism
D003838:Deoxyadenine Nucleotides; D046911:Macromolecular Substances; D000255:Adenosine Triphosphate; D012264:Ribonucleotide Reductases; C026600:2'-deoxyadenosine triphosphate