Regulation of T4 phage aerobic ribonucleotide reductase. Simultaneous assay of the four activities.
We have devised an assay procedure that permits simultaneous monitoring of the four activities of ribonucleotide reductase. Using this assay, we have compared the reduction of all four substrates by the T4 bacteriophage aerobic ribonucleotide reductase within different allosteric environments. Specifically, we compared the relative turnover rates by the enzyme when activated with "in vivo" concentrations of the known allosteric effectors versus activation by ATP alone. Consistent with the known allosteric properties of this enzyme, our results show that ATP does act as a general activator, although the rate of purine nucleotide reduction was approximately 5% of the rate for the pyrimidine nucleotides. However, addition of the allosteric effectors at their estimated physiological concentrations dramatically changed the relative rates of substrate reduction, creating a more "balanced" pool of products. Addition of the substrates at their respective in vivo concentrations further pushed rates of product formation toward a ratio similar to the base composition of the T4 genome. The similarity of the product profile produced under in vivo conditions to the genomic composition of T4 phage is discussed.
Hendricks
S P
SP
Department of Biochemistry and Biophysics, Oregon State University, Corvallis, Oregon 97331-7305, USA.
Mathews
C K
CK
D000255:Adenosine Triphosphate / Q000378:metabolism / Q000494:pharmacology; D000332:Aerobiosis; D000494:Allosteric Regulation; D017122:Bacteriophage T4 / Q000201:enzymology* / Q000235:genetics; D002846:Chromatography, Affinity; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D003854:Deoxyribonucleotides / Q000302:isolation & purification / Q000378:metabolism*; D004789:Enzyme Activation; D016679:Genome, Viral; D007700:Kinetics; D015203:Reproducibility of Results; D012264:Ribonucleotide Reductases / Q000737:chemistry / Q000378:metabolism*; D012265:Ribonucleotides / Q000302:isolation & purification / Q000378:metabolism*; D013379:Substrate Specificity
D003854:Deoxyribonucleotides; D012265:Ribonucleotides; D000255:Adenosine Triphosphate; D012264:Ribonucleotide Reductases