Herpes Simplex Virus-induced Ribonucleotide Reductase: Development of Antibodies Specific for the Enzyme

Abstract

The characterization of a novel ribonucleotide reductase induced in herpes simplex virus type 2 (HSV-2)-infected mammalian cells was reported. The virus-induced enzyme was partially purified, free of the constitutive cell isozyme, by fractionation of infected cell extracts with ammonium sulfate. A further purification of the virus-induced enzyme and the development of a rabbit antiserum capable of specifically inhibiting its activity is described. Enzymically active salt fractions from infected hamster kidney BHk-21 cell extracts were sedimented through glycerol gradients; the virus-induced enzyme sedimented .apprx. 2.5 times faster than the constitutive, or control, enzyme and was separated from the majority of the protein in the sample. Immunization of rabbits with the partially purified enzyme preparation recovered from gradients resulted in the synthesis of antibodies which completely and specifically inhibited the virus-induced reductase and precipitated 1 major polypeptide and a few minor species from both HSV-1 and HSV-2-infected cells. The antibodies exhibited much greater affinity for the HSV-2 than the HSV-1 antigen. Evidently, the virus-induced enzyme differs antigenically, as well as biochemically, from the constitutive cell isozyme. The hypothesis that the enzyme is virus-coded, is supported.