title: Curcumin Inhibits Protein Kinase Cα Activity by Binding to Its C1 Domain.
abstract: Curcumin is a polyphenolic nutraceutical that acts on multiple biological targets, including protein kinase C (PKC). PKC is a family of serine/threonine kinases central to intracellular signal transduction. We have recently shown that curcumin selectively inhibits PKCα, but not PKCε, in CHO-K1 cells [Pany, S. (2016) Biochemistry 55, 2135-2143]. To understand which domain(s) of PKCα is responsible for curcumin binding and inhibitory activity, we made several domain-swapped mutants in which the C1 (combination of C1A and C1B) and C2 domains are swapped between PKCα and PKCε. Phorbol ester-induced membrane translocation studies using confocal microscopy and immunoblotting revealed that curcumin inhibited phorbol ester-induced membrane translocation of PKCε mutants, in which the εC1 domain was replaced with αC1, but not the PKCα mutant in which αC1 was replaced with the εC1 domain, suggesting that αC1 is a determinant for curcumin's inhibitory effect. In addition, curcumin inhibited membrane translocation of PKCε mutants, in which the εC1A and εC1B domains were replaced with the αC1A and αC1B domains, respectively, indicating the role of both αC1A and αC1B domains in curcumin's inhibitory effects. Phorbol 13-acetate inhibited the binding of curcumin to αC1A and αC1B with IC50values of 6.27 and 4.47 μM, respectively. Molecular docking and molecular dynamics studies also supported the higher affinity of curcumin for αC1B than for αC1A. The C2 domain-swapped mutants were inactive in phorbol ester-induced membrane translocation. These results indicate that curcumin binds to the C1 domain of PKCα and highlight the importance of this domain in achieving PKC isoform selectivity.
authors: Department of Pharmacological and Pharmaceutical Sciences, College of Pharmacy, University of Houston , Houston, Texas 77204, United States.
authors: Satyabrata
authors: S
authors: Pany
authors: Department of Pharmacological and Pharmaceutical Sciences, College of Pharmacy, University of Houston , Houston, Texas 77204, United States.
authors: Youngki
authors: Y
authors: You
authors: Department of Pharmacological and Pharmaceutical Sciences, College of Pharmacy, University of Houston , Houston, Texas 77204, United States.
authors: Joydip
authors: J
authors: Das
mesh_terms: D001665:Binding Sites / Q000235:genetics; D001667:Binding, Competitive; D055162:Biocatalysis / Q000187:drug effects; D003474:Curcumin / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D057809:HEK293 Cells; D006801:Humans; D015151:Immunoblotting; D007700:Kinetics; D018613:Microscopy, Confocal; D056004:Molecular Dynamics Simulation; D009154:Mutation; D010703:Phorbol Esters / Q000494:pharmacology; D011485:Protein Binding; D000072417:Protein Domains; D051571:Protein Kinase C-alpha / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D051744:Protein Kinase C-epsilon / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D021381:Protein Transport / Q000187:drug effects / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D004791:Enzyme Inhibitors; D010703:Phorbol Esters; D011993:Recombinant Fusion Proteins; D051571:Protein Kinase C-alpha; D051744:Protein Kinase C-epsilon; D003474:Curcumin