THE R1 SUBUNIT OF HERPES-SIMPLEX VIRUS RIBONUCLEOTIDE REDUCTASE IS A GOOD SUBSTRATE FOR HOST-CELL PROTEIN-KINASES BUT IS NOT ITSELF A PROTEIN-KINASE

The N terminus of the R1 subunit of herpes simplex virus type 2 ribonu cleotide reductase is believed to be a protein kinase domain mainly be cause the R1 protein was phosphorylated in a protein kinase assay on b lot. Using Escherichia coil and adenovirus expression vectors to produ ce R1, we found that, whereas the reductase activity of both recombina nt proteins was similar, efficient phosphorylation of R1 and casein in the presence of Mg2+ was obtained only with the R1 purified from euka ryotic cells, Phosphorylation of this R1, in solution or on blot, resu lts mainly from the activity of casein kinase II (CKII), a co-purifyin g protein kinase. Labeling on blot occurs from CKII Leakage off the me mbrane and its subsequent high affinity binding to in vivo CKII-phosph orylated R1. CKII target sites were mapped to an acidic serine-rich se gment of the R1 N terminus. Improvement in purification of the R1 expr essed in eukaryotic cells nearly completely abolished its phosphorylat ion potential, An extremely low level of phosphorylation observed in t he presence of Mn2+ with the R1 produced in E. coli was probably due t o an unidentified prokaryotic protein kinase. These results provide ev idence that the herpes simplex virus type 2 R1 does not possess an int rinsic protein kinase activity.