Binding of ferulic acid to cytochrome c enhances stability of the protein at physiological pH and inhibits cytochrome c-induced apoptosis.
Ferulic acid (FA) is one of the most effective components of a traditional Chinese medicine, angelica, and cytochrome c plays a vital role in apoptosis. Here we report the application of fluorescence spectroscopy, isothermal titration calorimetry (ITC), differential scanning calorimetry (DSC), and circular dichroism (CD) to investigate the mechanism for the interaction of bovine heart cytochrome c with FA and the effect of the binding on native state stability of the protein at physiological pH. Fluorescence spectroscopic studies together with ITC measurements indicate that FA binds to cytochrome c with moderate affinity and quenches the intrinsic fluorescence of the protein in a static way. ITC experiments show that the interaction of cytochrome c with FA is driven by a moderately favorable entropy increase in combination with a less favorable enthalpy decrease for the first binding site of the protein. The melting temperature of cytochrome c in the presence of FA measured by DSC and CD increases 4.0 and 5.0 degrees C, respectively, compared with that in the absence of FA. Taken together, these results indicate that FA binds to and stabilizes cytochrome c at physiological pH. Furthermore, binding of FA to cytochrome c inhibits cytochrome c-induce apoptosis of human hepatoma cell line SMMC-7721. Our data provide insight into the mechanism of drug-protein interactions, and will be helpful to the understanding of the mechanism for FA-inhibited and cytochrome c-induced apoptosis.
Yang
Fang
F
State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan 430072, China.
Zhou
Bing-Rui
BR
Zhang
Peng
P
Zhao
Yan-Feng
YF
Chen
Jie
J
Liang
Yi
Y
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D002151:Calorimetry; D002417:Cattle; D002478:Cells, Cultured; D002942:Circular Dichroism; D003373:Coumaric Acids / Q000737:chemistry* / Q000494:pharmacology*; D045304:Cytochromes c / Q000737:chemistry* / Q000378:metabolism*; D004247:DNA / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D015394:Molecular Structure; D011485:Protein Binding; D013696:Temperature
D003373:Coumaric Acids; D045304:Cytochromes c; D004247:DNA; C004999:ferulic acid