DIMERIZATION OF THE HUMAN CYTOMEGALOVIRUS PROTEASE - KINETIC AND BIOCHEMICAL-CHARACTERIZATION OF THE CATALYTIC HOMODIMER

The single-chain 28 kDa human cytomegalovirus (HCMV) protease catalyti c domain containing the A143Q mutation has been kinetically and confor mationally characterized, The specific activity of the HCMV A143Q prot ease (HCMVp) increases as the protease concentration increases, sugges ting that this protease oligomerizes at high protein concentration to form a more active species, Both cross-linking and light-scattering st udies of HCMVp show the existence of a homodimer with an apparent mole cular mass of 56 kDa under low ionic strength and high protein concent ration. The cosolvent and solute effects of glycerol, trisodium citrat e, and NaCl as well as the temperature effects on the HCMVp activity a nd quaternary structure were investigated. The effects induced by coso lvents and temperature can largely be explained by their influences on the dimerization or oligomerization state nf HCMVp. The dissociation constant (K-d) for the HCMVp homodimer was determined to be 8 +/- 1 mu M with all activity attributed to the dimeric form. Monomeric HCMVp, is inactive. This report demonstrates that in vitro, HCMV A143Q protea se exists as an obligate catalytic homodimer, This protease dimerizati on may have regulatory significance during viral replication.