Analytical and Bioanalytical Chemistry

, Volume 408, Issue 18, pp 4935–4941 | Cite as

Redox-dependent interactions between reduced/oxidized cytochrome c and cytochrome c oxidase evaluated by in-situ electrochemical surface plasmon resonance

Research Paper

Abstract

The interactions between the redox couple of cytochrome c (Cyt c) and cytochrome c oxidase (COX) were investigated at a mimic redox-modulated interface by using an electrochemical surface plasmon resonance (EC-SPR) system. Although early studies of the binding between COX and Cyt c have been conducted using several techniques in homogeneous solutions, a problem still inherent is that ferro-cytochrome c (Cyt cred), the reduced form of Cyt c, can be easily oxidized into ferri-cytochrome c (Cyt cox) and adversely impact the accuracy and reproducibility of the binding measurements. In order to realize reliable redox-dependent binding tests, here the Cyt cred is quantitatively electro-generated from Cyt cox by in situ cathodic polarization in a flow cell. Then the kinetic and dissociation constants of the bindings between COX and Cyt cred/Cyt cox can be evaluated accurately. In this study, the values of association/dissociation rate constants (ka, kd) for both COX/Cyt cred and COX/Cyt cox were obtained. The dissociation constants, KD, were finally calculated as 3.33 × 10–8 mol · L–1 for COX/Cyt cred and 4.25 × 10–5 mol · L–1 for COX/Cyt cox, respectively. In-situ EC-SPR is promising for better mimicking the in vivo condition that COX is embedded in the inner mitochondrial membrane and Cyt c acts as an electron shuttle in the mobile phase. It is an effective method for the investigation of redox-dependent biomolecular interactions.

Graphical Abstract

Schematic representation of the experimental designs using EC-SPR system. (a) the Au-Cys-COX SPR chip with SAM layers. (b) redox-modulated Cyt c and its binding onto pre-immobilized COX

Keywords

Electrochemical surface plasmon resonance (EC-SPR) Cytochrome c (Cyt cCytochrome c oxidase (COX) Redox-dependent interaction Mimic interface Association/dissociation rate constant 

Supplementary material

216_2016_9586_MOESM1_ESM.pdf (476 kb)
ESM 1(PDF 476 kb)

References

  1. 1.
    Reed DE, Hawkridge FM. Direct electron transfer reactions of cytochrome c at silver electrodes. Anal Chem. 1987;59(19):2334–9.CrossRefGoogle Scholar
  2. 2.
    Hawkridge FM, Taniguchi I. The direct electron transfer reactions of cytochrome c at electrode surfaces. Comments Inorg Chem. 1995;17(3):163–87.CrossRefGoogle Scholar
  3. 3.
    Lee M-W, Park SC, Yang YG, Yim SO, Chae HS, Bach J-H, et al. The involvement of reactive oxygen species (ROS) and p38 mitogen-activated protein (MAP) kinase in TRAIL/Apo2L-induced apoptosis. FEBS Lett. 2002;512(1):313–8.CrossRefGoogle Scholar
  4. 4.
    Ferguson HA, Marietta PM, Van Den Berg CL. UV-induced apoptosis is mediated independent of caspase-9 in MCF-7 cells: a model for cytochrome c resistance. J Biol Chem. 2003;278(46):45793–800.CrossRefGoogle Scholar
  5. 5.
    Huttemann M, Pecina P, Rainbolt M, Sanderson TH, Kagan VE, Samavati L, et al. The multiple functions of cytochrome c and their regulation in life and death decisions of the mammalian cell: from respiration to apoptosis. Mitochondrion. 2011;11(3):369–81.CrossRefGoogle Scholar
  6. 6.
    Michel H. Cytochrome c oxidase: catalytic cycle and mechanisms of proton pumping—a discussion. Biochemistry. 1999;38(46):15129–40.CrossRefGoogle Scholar
  7. 7.
    Nelson DL, Cox MM. Lehninger principles of biochemistry. Freeman, New York; 2004.Google Scholar
  8. 8.
    Speck SH, Dye D, Margoliash E. Single catalytic site model for the oxidation of ferrocytochrome c by mitochondrial cytochrome c oxidase. Proc Natl Acad Sci U S A. 1984;81(2):347–51.CrossRefGoogle Scholar
  9. 9.
    Michel B, Bosshard HR. Oxidation of cytochrome c by cytochrome c oxidase: spectroscopic binding studies and steady-state kinetics support a conformational transition mechanism. Biochemistry. 1989;28(1):244–52.CrossRefGoogle Scholar
  10. 10.
    Wilson MT, Greenwood C, Brunori M, Antonini E. Kinetic studies on the reaction between cytochrome c oxidase and ferrocytochrome c. Biochem J. 1975;147(1):145–53.CrossRefGoogle Scholar
  11. 11.
    Ferguson-Miller S, Brautigan DL, Margoliash E. Definition of cytochrome c binding domains by chemical modification. III. Kinetics of reaction of carboxydinitrophenyl cytochrome c with cytochrome c oxidase. J Biol Chem. 1978;253(1):149–59.Google Scholar
  12. 12.
    Green RJ, Frazier RA, Shakesheff KM, Davies MC, Roberts CJ, Tendler SJ. Surface plasmon resonance analysis of dynamic biological interactions with biomaterials. Biomaterials. 2000;21(18):1823–35.CrossRefGoogle Scholar
  13. 13.
    Malmqvist M. Biospecific interaction analysis using biosensor technology. Nat Int Wkly J Sci. 1993;361(6408):186–7.Google Scholar
  14. 14.
    Karlsson R. SPR for molecular interaction analysis: a review of emerging application areas. J Mol Recognit. 2004;17(3):151–61.CrossRefGoogle Scholar
  15. 15.
    Jiang Z, Yun Q, Zhen P, Chen S, Shu C, Deng C, et al. The simultaneous detection of free and total prostate antigen in serum samples with high sensitivity and specificity by using the dual-channel surface plasmon resonance. Biosens Bioelectron. 2014;62(20):268–73.CrossRefGoogle Scholar
  16. 16.
    Iwasaki Y, Horiuchi T, Morita M, Niwa O. Analysis of electrochemical processes using surface plasmon resonance. Sensors Actuators B. 1998;50(2):145–8.CrossRefGoogle Scholar
  17. 17.
    Iwasaki Y, Horiuchi T, Niwa O. Detection of electrochemical enzymatic reactions by surface plasmon resonance measurement. Anal Chem. 2001;73(7):1595–8.CrossRefGoogle Scholar
  18. 18.
    Hua D, Cao X, Chang ML, Hu W. An in situ electrochemical surface plasmon resonance immunosensor with polypyrrole propylic acid film: comparison between SPR and electrochemical responses from polymer formation to protein immunosensing. Biosens Bioelectron. 2008;23(7):1055–62.CrossRefGoogle Scholar
  19. 19.
    Shaopeng W, Forzani ES, Nongjian T. Detection of heavy metal ions in water by high-resolution surface plasmon resonance spectroscopy combined with anodic stripping voltammetry. Anal Chem. 2007;79(12):4427–32.CrossRefGoogle Scholar
  20. 20.
    Yuting H, Ningning X, Shu C, Chunyan D, Juan X. Controllable release and high‐efficiency collection of hydrogen peroxide: application on the quantitative investigation of biomolecule oxidation induced by reactive oxygen species. Electroanalysis. 2014;26(7):1497–503.CrossRefGoogle Scholar
  21. 21.
    Yao X, Yang M, Wang Y, Hu Z. Study of the ferrocenylalkanethiol self-assembled monolayers by electrochemical surface plasmon resonance. Sensors Actuators B. 2007;122(2):351–6.CrossRefGoogle Scholar
  22. 22.
    Boussaad S, Pean J, Tao NJ. High-resolution multiwavelength surface plasmon resonance spectroscopy for probing conformational and electronic changes in redox proteins. Anal Chem. 2000;72(1):222–6.CrossRefGoogle Scholar
  23. 23.
    Juan X, Jun G, Feimeng Z. Scanning electrochemical microscopy combined with surface plasmon resonance: studies of localized film thickness variations and molecular conformation changes. Anal Chem. 2006;78(5):1418–23.CrossRefGoogle Scholar
  24. 24.
    Zhai P, Guo J, Xiang J, Zhou F. Electrochemical surface plasmon resonance spectroscopy at bilayered silver/gold films. J Phys Chem C. 2007;111(2):981–6.CrossRefGoogle Scholar
  25. 25.
    Murgida DH, Hildebrandt P. Electron-transfer processes of cytochrome c at interfaces. New insights by surface-enhanced resonance Raman spectroscopy. Acc Chem Res. 2004;37(11):854–61.CrossRefGoogle Scholar
  26. 26.
    Fedurco M. Redox reactions of heme-containing metalloproteins: dynamic effects of self-assembled monolayers on thermodynamics and kinetics of cytochrome c electron-transfer reactions. Coord Chem Rev. 2000;209:263–331.CrossRefGoogle Scholar
  27. 27.
    Liu Y-C, Cui S-Q, Zhao J, Yang Z-S. Direct electrochemistry behavior of cytochrome c/l-cysteine modified electrode and its electrocatalytic oxidation to nitric oxide. Bioelectrochemistry. 2007;70(2):416–20.CrossRefGoogle Scholar
  28. 28.
    Di Gleria K, Hill HAO, Lowe VJ, Page DJ. Direct electrochemistry of horse-heart cytochrome c at amino acid-modified gold electrodes. J Electroanal Chem Interfacial Electrochem. 1986;213(2):333–8.CrossRefGoogle Scholar
  29. 29.
    Zhang Y, Xu M, Wang Y, Toledo F, Zhou F. Studies of metal ion binding by apo-metallothioneins attached onto preformed self-assembled monolayers using a highly sensitive surface plasmon resonance spectrometer. Sensors Actuators B. 2007;123(2):784–92.CrossRefGoogle Scholar
  30. 30.
    Bin Y, Chen S, Xiang J. pH-dependent kinetics of copper ions binding to amyloid-β peptide. J Inorg Biochem. 2013;119:21–7.CrossRefGoogle Scholar
  31. 31.
    Oshannessy DJ, Brighamburke M, Soneson KK, Hensley P, Brooks I. Determination of rate and equilibrium binding constants for macromolecular interactions using surface plasmon resonance: use of nonlinear least squares analysis methods. Anal Biochem. 1993;212(2):457–68.Google Scholar
  32. 32.
    Adamson AW. Physical chemistry of surfaces. 6th ed. Wiley: New York; 1997.Google Scholar
  33. 33.
    Ferguson-Miller S, Brautigan DL, Margoliash E. Correlation of the kinetics of electron transfer activity of various eukaryotic cytochromes c with binding to mitochondrial cytochrome c oxidase. J Biol Chem. 1976;251(4):1104–15.Google Scholar
  34. 34.
    Rieder R, Bosshard HR. The cytochrome c oxidase binding site on cytochrome c. Differential chemical modification of lysine residues in free and oxidase-bound cytochrome c. J Biol Chem. 1978;253(17):6045–53.Google Scholar
  35. 35.
    Osheroff N, Brautigan DL, Margoliash E. Definition of enzymic interaction domains on cytochrome c. Purification and activity of singly substituted carboxydinitrophenyl-lysine 7, 25, 73, 86, and 99 cytochromes c. J Biol Chem. 1980;255(17):8245–51.Google Scholar
  36. 36.
    Michel B, Bosshard HR. Spectroscopic analysis of the interaction between cytochrome c and cytochrome c oxidase. J Biol Chem. 1984;259(16):10085–91.Google Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2016

Authors and Affiliations

  1. 1.College of Chemistry and Chemical EngineeringCentral South UniversityChangshaChina

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