title: Active human cytomegalovirus protease is a dimer.
abstract: The quaternary state of the human cytomegalovirus (hCMV) protease has been analyzed in relation to its catalysis of peptide hydrolysis. Based on results obtained from steady state kinetics, size exclusion chromatography, and velocity sedimentation, the hCMV protease exists in a monomer-dimer equilibrium. Dimerization of the protease is enhanced by the presence of glycerol and high concentrations of enzyme. Isolation of monomeric and dimeric species eluted from a size exclusion column, followed by immediate assay, identifies the dimer as the active species. Activity measurements conducted with a range of enzyme concentrations are also consistent with a kinetic model in which only the dimeric hCMV protease is active. Using this model, the dissociation constant of the protease is 6.6 microM in 10% glycerol and 0.55 microM in 20% glycerol at 30 degrees C and pH 7.5.
authors.affiliation: Department of Antiviral Research, Merck Research Laboratories, West Point, Pennsylvania 19486, USA.
authors.forename: P L
authors.initials: PL
authors.lastname: Darke
authors.forename: J L
authors.initials: JL
authors.lastname: Cole
authors.forename: L
authors.initials: L
authors.lastname: Waxman
authors.forename: D L
authors.initials: DL
authors.lastname: Hall
authors.forename: M K
authors.initials: MK
authors.lastname: Sardana
authors.forename: L C
authors.initials: LC
authors.lastname: Kuo
mesh_terms: D000595:Amino Acid Sequence; D002850:Chromatography, Gel; D003587:Cytomegalovirus / Q000201:enzymology*; D010450:Endopeptidases / Q000096:biosynthesis / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D005990:Glycerol / Q000494:pharmacology; D006801:Humans; D007700:Kinetics; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009842:Oligopeptides / Q000138:chemical synthesis / Q000378:metabolism; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D012697:Serine Endopeptidases; D013379:Substrate Specificity
chemical_list: D046911:Macromolecular Substances; D009842:Oligopeptides; D011994:Recombinant Proteins; D010450:Endopeptidases; D012697:Serine Endopeptidases; C071910:assemblin; D005990:Glycerol