Direct electron injection from electrodes to cytochrome P450_cam in biomembrane-like films

Abstract

Direct, reversible, electron transfer has been demonstrated between electrodes and native cytochrome P450_cam (cyt P450_cam) in lipid films. Electron injection directly from electrodes into the haem Fe^III of cyt P450_cam has been confirmed by the influence of the known reaction of cyt P450_camFe^II with CO on the voltammetric midpoint potential. Analysis of square wave voltammograms suggested a distribution of enzyme formal potentials in the films. With oxygen present, more than one electron was injected into the enzyme in films, mimicking in vivo electron acceptance by cyt P450Fe^III and cyt P450Fe^II–O₂ during catalytic oxidations. Cyt P450_cam films also catalysed electrochemically driven reduction of trichloroacetic acid in anaerobic solutions. These stable enzyme–lipid films may find applications in fundamental biochemical and toxicity studies, biocatalysis and biosensors.

References

1. Cytochrome P450, ed. P. R. Ortiz de Montellano, Plenum, New York, 1986.
2. Cytochrome P450, ed. J. B. Schenkman and H. Greim, Springer-Verlag, Berlin, 1993.
3. F. P. Guengerich and T. L. MacDonald, Acc. Chem. Res., 1984, 17, 9.
4. Cytochrome P450, ed. T. Omura, Y. Ishimura and Y. Fujii-Kuriyama, 2nd edn., Kodansha VCH, Tokyo, 1993.
5. C. E. Castro, R. S. Wade and N. O. Belser, Biochemistry, 1985, 24, 204.
6. S. Li and L. P. Wackett, Biochemistry, 1993, 32, 9355.
7. G. S. Koe and V. L. Vilker, Biotechnol. Prog., 1993, 9, 608.
8. (a) F. A. Armstrong, H. A. O. Hill and N. J. Walton, Acc. Chem. Res., 1988, 21, 407; (b) F. A. Armstrong, in Bioinorganic Chemistry, Structure and Bonding 72, Springer-Verlag, Berlin, 1990, pp. 137–221.
9. A. Heller, Acc. Chem. Res., 1990, 23, 128.
10. (a) M. J. Tarlov and E. F. Bowden, J. Am. Chem. Soc., 1991, 113, 1847; (b) E. F. Bowden, F. M. Hawkridge and H. N. Blount, in Comprehensive Treatise of Electrochem., ed. S. Srinivasan, Y. A. Chizmadzhev, J. O'M. Bockris, B. E. Conway and E. Yeager, Plenum, New York, 1985, vol. 10, pp. 297–346.
11. (a) J. F. Rusling and A.-E. F. Nassar, J. Am. Chem. Soc., 1993, 115, 11891; (b) A.-E. F. Nassar, Y. Narikiyo, T. Sagara, N. Nakashima and J. F. Rusling, J. Chem. Soc., Faraday Trans., 1995, 91, 1775.
12. A.-E. F. Nassar, Z. Zhang, V. Chynwat, H. A. Frank, J. F. Rusling and K. Suga, J. Phys. Chem., 1995, 99, 11013.
13. (a) P. Bianco and J. Haladjian, J. Electroanal. Chem., 1994, 367, 79; (b) P. Bianco and J. Haladjian, Electrochim. Acta, 1994, 39, 911; (c) J. Hanzlik, P. Bianco and J. Haladjian, J. Electroanal. Chem., 1995, 380, 287; (d) P. Bianco and J. Haladjian, Electroanalysis, 1995, 7, 442; (e) M. Tominaga, J. Yanagimoto, A.-E. F. Nassar, J. F. Rusling and N. Nakashima, Chem. Lett., 1996, 523; (f) Z. Lu, Q. Huang and J. F. Rusling, J. Electroanal. Chem., 1996, in press; (g) Q. Huang, Z. Lu and J. F. Rusling, Langmuir, 1996, 12, 5472.
14. J. Kazlauskaite, A. C. G. Westlake, L.-L. Wong and H. A. O. Hill, J. Chem. Soc., Chem. Commun., 1996, 2189.
15. (a) F. Scheller, R. Renneberg, G. Strnad, K. Pommerening and P. Mohr, Bioelectrochem. Bioenerg., 1977, 4, 500; (b) A. I. Archakov, B. A. Kuznetsov, M. V. Izotov and I. I. Karuzina, Dokl. Akad. Nauk. SSSR, 1981, 258, 216.
16. V. L. Vilker, F. Kahn, D. Shen, M. M. Baizer and K. Nobe, in Redox Chemistry and Interfacial Behavior of Biological Molecules, ed. G. Dryhurst and K. Niki, Plenum, New York, 1988, pp. 105–112.
17. K. M. Faulkner, M. S. Shet, C. W. Fisher and R. W. Estabrook, Proc. Natl. Acad. Sci., USA, 1995, 92, 7705.
18. D. H. O'Keefe, R. E. Ebel and J. A. Peterson, Methods Enzymol., 1978, 52, 151.
19. R. W. Murray, in Electroanalytical Chemistry, ed. A. J. Bard, Marcel Dekker, New York, 1984, vol. 13, pp. 191–368.
20. F. P. Guengerich, D. P. Ballou and M. J. Coon, J. Biol. Chem., 1975, 250, 7405.
21. J. A. Peterson and B. W. Griffin, Arch. Biochem. Biophys., 1972, 151, 427.
22. J. A. Peterson, Arch. Biochem. Biophys., 1971, 144, 678.
23. A. J. Bard and L. R. Faulkner, Electrochemical Methods, Wiley, New York, 1980.
24. E. Laviron, J. Electroanal. Chem., 1979, 101, 19.
25. J. Osteryoung and J. J. O'Dea, in Electroanalytical Chemistry, ed. A. J. Bard, Marcel Dekker, New York, 1986, vol. 14, pp. 209–308.
26. J. J. O'Dea and J. Osteryoung, Anal. Chem., 1993, 65, 3090.
27. (a) M. S. Wrighton, M. C. Pallazzotto, A. B. Bocarsly, J. M. Bolts, A. B. Fischer and L. Nadjo, J. Am. Chem. Soc., 1978, 100, 7264; (b) P. J. Peerce and A. J. Bard, J. Electroanal. Chem., 1980, 114, 89; (c) W. J. Albery, M. G. Boutelle, P. J. Colby and A. R. Hillman, J. Electroanal. Chem., 1982, 133, 135; (d) G. K. Rowe, M. T. Carter, J. N. Richardson and R. W. Murray, Langmuir, 1995, 11, 1797; (e) E. Sabatani and F. C. Anson, J. Electroanal. Chem., 1995, 386, 111; T. M. Nahir and E. F. Bowden, J. Electroanal. Chem., 1996, 410, 9.
28. Z. Zhang and J. F. Rusling, Biophys. Chem., 1996, in press.
29. J. F. Rusling and T. F. Kumosinski, Nonlinear Computer Modeling of Chemical and Biochemical Data, Academic Press, New York, 1996, pp. 177–205.
30. J. F. Rusling, Acc. Chem. Res., 1991, 24, 75.
31. A.-E. F. Nassar, J. M. Bobbitt, J. D. Stuart and J. F. Rusling, J. Am. Chem. Soc., 1995, 117, 10986.
32. J. F. Rusling, Microporous Materials, 1994, 3, 1.
33. S. Manne and H. E. Gaub, Science, 1995, 270, 1480.
34. S. G. Sligar, D. L. Cinti, G. G. Gibson and J. B. Schenkman, Biochem. Biophys. Res. Commun., 1979, 90, 925.
35. N. C. Veitch and R. P. J. Williams, in Relationships Between Structure and Functions of Cytochrome P450, ed. K. Ruckpaul and H. Rein, Academie Verlag, Berlin, 1992, pp. 279–320.
36. J. D. Lipscomb, S. G. Sligar, M. J. Namtvedt and I. C. Gunsalus, J. Biol. Chem., 1976, 251, 1116.