ALLOSTERIC EFFECTORS ARE REQUIRED FOR SUBUNIT ASSOCIATION IN T4 PHAGERIBONUCLEOTIDE REDUCTASE

Bacteriophage T4 encodes its own aerobic ribonucleotide reductase (RNR ), which reduces ribonucleoside diphosphates to the corresponding deox yribonucleoside diphosphates. T4 RNR is composed of homodimeric large (R1) and small (R2) subunits. Intricate regulation of enzymatic activi ty is accomplished by the binding of nucleotide effecters to R1. Bergl und (Berglund, O. (1972) J. Biol. Chem. 247, 7270-7275) described simi larities between T4 RNR and the corresponding enzyme from aerobic Esch erichia coli. An important difference, however, is that T4 RNR forms a tight R1.R2 complex, while the E. coli R1 and R2 more readily dissoci ate. In this study we purified the phage R2 subunit from an overexpres sion vector constructed by Tseng et al. (Tseng, M., Hilfinger, J., He, P., and Greenberg, R. (1992) J. Bacteriol. 174, 5740-5744) and used t his as an immunogen to generate polyclonal antiserum. Using co-immunop recipitation techniques, we probed in vitro for interactions between t he phage-induced R1 and R2 subunits. Our studies indicate that tight b inding of the phage RNR subunits is completely dependent upon the know n allosteric effecters of the enzyme. Once the R1.R2 holoenzyme has be en formed it appears to be remarkably stable when in the presence of d ATP. However, if dATP is removed, the R1.R2 complex readily dissociate s.