Hepatitis C virus NS3 protease is activated by low concentrations of protease inhibitors
2009 (English)In: Biochemistry, ISSN 0006-2960, E-ISSN 1520-4995, Vol. 48, no 48, 11592-11602 p.Article in journal (Refereed) Published
The nonstructural protein 3 (NS3) of hepatitis C virus (HCV) is a bifunctional enzyme with a protease and a helicase functionality located in each of the two domains of the single peptide chain. There is little experimental evidence for a functional role of this unexpected arrangement since artificial single domain forms of both enzymes are catalytically competent. We have observed that low concentrations of certain protease inhibitors activate the protease of full-length NS3 from HCV genotype 1a with up to 100%, depending on the preincubation time and the inhibitor used. The activation was reduced, but not eliminated, by increased ionic strength, lowered glycerol concentration, or lowered pH. In all cases, it was at the expense of a significant loss of activity. Activation was not seen with the artificial protease domain of genotype 1b NS3 fused with a fragment of the NS4A cofactor. This truncated and covalently modified enzyme form was much less active and exhibited fundamentally different catalytic properties to the full-length NS3 protease without the fused cofactor. The most plausible explanation for the activation was found to involve a slow transition between two enzyme conformations, which differed in their catalytic ability and affinity for inhibitors. Equations derived based on this assumption resulted in better fits to the experimental data than the equation for simple competitive inhibition. The mechanism may involve an inhibitor-induced stabilization of the helicase domain in a conformation that enhances the protease activity, or an improved alignment of the catalytic triad in the protease. The proposed mnemonic mechanism and derived equations are viable for both these explanations and can serve as a basic framework for future studies of enzymes activated by inhibitors or other ligands.
Place, publisher, year, edition, pages
2009. Vol. 48, no 48, 11592-11602 p.
Biocatalysis, Enzyme Activation, Enzyme Activation: drug effects, Genotype, Glycerol, Glycerol: chemistry, Hydrogen-Ion Concentration, Osmolar Concentration, Protease Inhibitors, Protease Inhibitors: pharmacology, Protein Conformation, Viral Nonstructural Proteins, Viral Nonstructural Proteins: antagonists & inhibitors, Viral Nonstructural Proteins: chemistry, Viral Nonstructural Proteins: metabolism
IdentifiersURN: urn:nbn:se:kth:diva-53380DOI: 10.1021/bi9016928ISI: 000272083900028PubMedID: 19839643OAI: oai:DiVA.org:kth-53380DiVA: diva2:470024
QC 201112282011-12-282011-12-282011-12-28Bibliographically approved