Dimerization and activation of the herpes simplex virus type 1 protease.
The quaternary state of the herpes simplex virus type 1 (HSV-1) protease has been analyzed in relation to its catalytic activity. The dependence of specific activity upon enzyme concentration indicated that association of the 27-kDa subunits strongly increased activity. Size-exclusion chromatography identified the association as a monomer-dimer equilibrium. Isolation of monomeric and dimeric species from a size-exclusion column followed by immediate assay identified the dimer as the active form of the enzyme. Activation of the protease by antichaotropic cosolvents correlated with changes in the monomer-dimer equilibrium. Thus, dimerization of the enzyme was enhanced in solvents containing glycerol or the anions citrate or phosphate. These are substances previously identified as activators of HSV-1 protease (Hall, D. L., and Darke, P. L. (1995) J. Biol. Chem. 270, 22697-22700). The relative potencies of these cosolvents as enzyme activators correlated with their efficiency in promoting dimerization. Under all solvent conditions examined, the dependence of specific activity upon enzyme concentration was consistent with a kinetic model in which only the dimer is active. Dissociation constants for the HSV-1 protease dimer determined with this model at 15 degrees C, pH 7.5, were 964 and 225 nM in 20% glycerol with 0.2 and 0.5 M citrate present, respectively. The activation of the HSV-1 protease by antichaotropic cosolvents was hereby shown to be similar in nature to the activation of the other well characterized herpesvirus protease, that from human cytomegalovirus.
Schmidt
U
U
Department of Antiviral Research, Merck Research Laboratories, West Point, Pennsylvania 19486, USA.
Darke
P L
PL
D000595:Amino Acid Sequence; D001704:Biopolymers; D002213:Capsid / Q000378:metabolism*; D002850:Chromatography, Gel; D004789:Enzyme Activation; D007700:Kinetics; D008969:Molecular Sequence Data; D011487:Protein Conformation; D012697:Serine Endopeptidases / Q000378:metabolism*; D014764:Viral Proteins
D001704:Biopolymers; D014764:Viral Proteins; C079287:scaffold protein, Herpes simplex virus-1; D012697:Serine Endopeptidases