Herpes simplex virus ribonucleotide reductase induced in infected BHK-21/C13 cells: biochemical evidence for the existence of two non-identical subunits, H1 and H2.
In nearly all systems studied, ribonucleotide reductase consists of two non-identical subunits. We present here the results of our study on herpes simplex virus (HSV) ribonucleotide reductase in favour of the existence of two subunits, H1 and H2, different from the mammalian subunits, M1 and M2. First, although the viral subunits could not be separated by Blue Sepharose chromatography (unlike mammalian subunits), they seemed to dissociate at very low protein concentration as suggested by the non-linear relationship between activity and low protein concentration. Second, pyridoxal phosphate (Pyr.P)-NaBH4 treatment and 4-methyl-5-amino-1-formylisoquinoline thiosemicarbazone (MAIQ) treatment of partially purified extract of mammalian ribonucleotide reductase which inactivated M1 and M2 respectively also inhibited the HSV ribonucleotide reductase. This activity could be restored by mixing Pyr.P-NaBH4-treated extracts with MAIQ-treated extracts of viral ribonucleotide reductase, suggesting that each treated extract contains one active subunit. Moreover, the addition of exogenous M1 or M2 subunits to one or the other of these two treated extracts did not produce any detectable reductase activity. Our interpretation of these results is that the two subunits H1 and H2 which could dissociate upon treatment did not form enzymically active hybrids with the mammalian subunits. Also, the higher degree of resistance to heat inactivation and to hydroxyurea of the viral reductase as compared to the mammalian enzyme suggests that H1 differs from M1 and H2 from M2.
Cohen
E A
EA
Charron
J
J
Perret
J
J
Langelier
Y
Y
D000818:Animals; D002478:Cells, Cultured; D002847:Chromatography, Agarose; D006224:Cricetinae; D004790:Enzyme Induction; D006358:Hot Temperature; D007546:Isoquinolines / Q000494:pharmacology; D011732:Pyridoxal Phosphate / Q000494:pharmacology; D012264:Ribonucleotide Reductases / Q000032:analysis* / Q000037:antagonists & inhibitors / Q000096:biosynthesis; D012685:Sepharose / Q000031:analogs & derivatives; D018139:Simplexvirus / Q000201:enzymology*
D007546:Isoquinolines; C013118:4-methyl-5-amino-1-formylisoquinoline thiosemicarbazone; D011732:Pyridoxal Phosphate; C029483:Blue-Sepharose; D012685:Sepharose; D012264:Ribonucleotide Reductases