Aurintricarboxylic acid modulates the affinity of hepatitis C virus NS3 helicase for both nucleic acid and ATP.
Aurintricarboxylic acid (ATA) is a potent inhibitor of many enzymes needed for cell and virus replication, such as polymerases, helicases, nucleases, and topoisomerases. This study examines how ATA interacts with the helicase encoded by the hepatitis C virus (HCV) and reveals that ATA interferes with both nucleic acid and ATP binding to the enzyme. We show that ATA directly binds HCV helicase to prevent the enzyme from interacting with nucleic acids and to modulate the affinity of HCV helicase for ATP, the fuel for helicase action. Amino acid substitutions in the helicase DNA binding cleft or its ATP binding site alter the ability of ATA to disrupt helicase-DNA interactions. These data, along with molecular modeling results, support the notion that an ATA polymer binds between Arg467 and Glu493 to prevent the helicase from binding either ATP or nucleic acids. We also characterize how ATA affects the kinetics of helicase-catalyzed ATP hydrolysis, and thermodynamic parameters describing the direct interaction between HCV helicase and ATA using microcalorimetry. The thermodynamics of ATA binding to HCV helicase reveal that ATA binding does not mimic nucleic acid binding in that ATA binding is driven by a smaller enthalpy change and an increase in entropy.
Department of Chemistry and Biochemistry, University of Wisconsin-Milwaukee , 3210 North Cramer Street, Milwaukee, Wisconsin 53211, United States.
William R
WR
Shadrick
Sourav
S
Mukherjee
Alicia M
AM
Hanson
Noreena L
NL
Sweeney
David N
DN
Frick
D000255:Adenosine Triphosphate / Q000378:metabolism*; D019943:Amino Acid Substitution; D001312:Aurintricarboxylic Acid / Q000494:pharmacology*; D002152:Calorimetry, Differential Scanning; D004247:DNA / Q000378:metabolism*; D016174:Hepacivirus / Q000201:enzymology; D008958:Models, Molecular; D017361:Viral Nonstructural Proteins / Q000037:antagonists & inhibitors / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases
D000255:Adenosine Triphosphate; D001312:Aurintricarboxylic Acid; D004247:DNA; D017361:Viral Nonstructural Proteins; C084422:NS3 protein, hepatitis C virus; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D053487:DEAD-box RNA Helicases; D000085242:Viral Proteases