A fluorescence imaging technique was used to monitor intracellular localization of protein kinase C (PKC) in U-87 MG human glioma cells in the presence of hypericin (Hyp) and phorbol 12-myristate-13-acetate (PMA). It is shown that PKC localization, which reflects its activity, is influenced by Hyp and this influence is different from that observed for PMA which acts as PKC activator. Fluorescence binding experiments were used to determine the binding constants of Hyp to several isoforms of PKC. The obtained values of Kds (∼100 nM) suggest that Hyp binds with high affinity to PKC. Finally, molecular modeling was used to compare structural models of the interaction of C1B domain of PKC (PKC isoforms α, δ, γ) with Hyp and our previously published model of the (C1B domain PKCγ)/PMA complex. The influence of Hyp on PKC translocation in U-87 MG cells in comparison with PMA, colocalization fluorescence pattern of Hyp and PKC, the higher binding affinity of Hyp to PKC in comparison with known binding constants of phorbol esters, as well as the binding mode of Hyp and PMA to the C1B domain of PKC suggested by molecular modeling, support the idea that Hyp and PMA might competitively bind to the regulatory domain of PKC.
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1 May 2006
Characterization of the Interaction of Hypericin with Protein Kinase C in U-87 MG Human Glioma Cells
Silvia Kocanova,
Tekla Hornakova,
Jozef Hritz,
Daniel Jancura,
Dusan Chorvat,
Anton Mateasik,
Jozef Ulicny,
Matthieu Refregiers,
Jean-Claude Maurizot,
Pavol Miskovsky
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Photochemistry and Photobiology
Vol. 82 • No. 3
May 2006
Vol. 82 • No. 3
May 2006