Photosynthesis Research

, Volume 42, Issue 2, pp 111–120 | Cite as

Effects of pH on the kinetics of redox reactions in and around the cytochromebf complex in an isolated system

  • A. B. Hope
  • P. Valente
  • D. B. Matthews
Regular Paper


Rate-coefficients describing the electron transfer reactions between P700 and plastocyanin, between cytochromef in cytochromebf complexes and plastocyanin, and between decyl plastoquinol and cytochromebf complexes were determined as a function of pH in the range 4–10 from flash-induced absorbancy changes at four wavelengths. The reactions between P700 and plastocyanin, and between cytochromef and plastocyanin were optimised when there was electrostatic interaction between ionised acidic groups in plastocyanin with a pKa of 4.3–4.7 and ionised basic constituents in P700 (assumed to be in the PSI-F subunit) and in cytochromef, with a pKb of 8.9–9.4. The basic groups are thought to be lysine rather than arginine. This mechanism agrees with that inferred from effects of ionic strength changes on rate-coefficients. The relation between the second-order rate-coefficient for decyl plastoquinol oxidation by thebf complex and pH was characterised by a pKa of 6.1. This is interpreted as showing that the anion radical form of that quinol, which has a pKa of 6, and which becomes progressively protonated when pH is changed from 7 to 5, is essential to reduce cytochromeb-563 (low potential) during quinol oxidation. Above pH 9, permanent effects were observed on this rate-coefficient, which were absent in the reactions between P700, plastocyanin and cytochromef.

Key words

electron transfers decyl plastoquinol plastocyanin P700 



anthraquinone sulphonate


3-(cyclohexylamino) propanesulphonic acid


cyclohexylaminoethanesulphonic acid






mid-point reduction potential


Rieske centre


high potential cytb-563


low potential cytb-563




2-(N-morpholino) ethanesulphonic acid


3-(N-morpholino)propanesulphonic acid










reaction centre of Photosystem I




Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Adam Z and Malkin R (1989) On the interaction between cytochromef and plastocyanin. Biochim Biophys Acta 975: 158–163Google Scholar
  2. Anderson GP, Draheim JE and Gross EL (1985) Plastocyanin conformation: The effect of the oxidation state on the pKa of nitrotyrosine-83. Biochim Biophys Acta 810: 123–131Google Scholar
  3. Bendall DS (1982) Photosynthetic cytochromes of oxygenic organisms. Biochim Biophys Acta 683: 119–151Google Scholar
  4. Beoku-Betts D, Chapman SK, Knox CV and Sykes AG (1985) Kinetic studies on 1:1 electron-transfer reactions involving blue copper proteins. 11. Effects of pH, competitive inhibitors and Chromium (III) modification on the reaction of plastocyanin with cytochromef. Inorg Chem 24: 1677–1681Google Scholar
  5. Bottin H and Mathis P (1985) Interaction of plastocyanin with the Photosystem I reaction center: a kinetic study by flash absorption spectroscopy. Biochemistry 24: 6453–6460Google Scholar
  6. Bruhn H, Nigam S and Holzwarth JF (1982) Catalytic influence of the environment on outer-sphere eletron-transfer reactions in aqueous solutions. Farad Disc Chem Soc 74: 129–140Google Scholar
  7. Burkey KO and Gross EL (1982) Chemical modification of spinach plastocyanin. Separation and characterisation of four different forms. Biochemistry 21: 5886–5890Google Scholar
  8. Christensen HEM, Conrad LS and Ulstrup J (1992) Effects of NO2-modification of Tyr83 on the reactivity of spinach plastocyanin with cytochromef. Biochim Biophys Acta 1099: 35–44Google Scholar
  9. Davidson E, Ohnishi T, Atta-Asafo-Adjei E and Daldal F (1992) Potential ligands to the [2Fe-2S] Rieske cluster of the cytochromebc 1 complex of Rhodobacter capsulata probed by site-directed mutagenesis. Biochemistry 31: 3342–3351Google Scholar
  10. Fersht AR (1993) Protein folding and stability: the pathway of folding of barnase. FEBS Lett 325: 5–16Google Scholar
  11. Gross EL (1993) Plastocyanin: Structure and function. Photosynth Res 37: 103–116Google Scholar
  12. Gross EL and Curtiss A (1991) The interaction of nitrotyrosine-83 plastocyanin with cytochromes f and c: pH dependence and the effect of an additional negative charge on plastocyanin. Biochim Biophys Acta 1056: 166–172Google Scholar
  13. Gross EL, Curtiss A, Durell SR and White D (1990) Chemical modification of spinach plastocyanin using 4-chloro-3,5-dinitrobenzoic acid: Characterisation of four singly-charged forms. Biochim Biophys Acta 1016: 107–114Google Scholar
  14. Guss JM and Freeman HC (1983) Structure of oxidised poplar plastocyanin at 1.6 Å resolution. J Mol Biol 169: 521–563Google Scholar
  15. Hauska G, Hurt E, Gabellini N, and Lockau W (1983) Comparative aspects of quinol-cytochromec/plastocyanin oxidoreductases. Biochim Biophys Acta 726: 97–133Google Scholar
  16. He S, Modi S, Bendall DS and Gray JC (1991) The surface-exposed tyrosine residue Tyr83 of pea plastocyanin in involved in both binding and electron transfer reactions with cytochromef. The EMBO J 10: 4011–4016Google Scholar
  17. Hippler M, Ratajczak R and Haehnel W (1989) Identification of the plastocyanin binding subunit of Photosystem I. FEBS Lett 250: 280–284Google Scholar
  18. Hope AB (1993) The chloroplast cytochromebf complex: A critical focus on function. Biochim Biophys Acta 1143: 1–22Google Scholar
  19. Hope AB, Huilgol RR, Panizzia M, Thompson M and Matthews DB (1992) The flash-induced turnover of cytochromeb-563, cytochromef and plastocyanin in chloroplasts. Models and estimation of kinetic parameters. Biochim Biophys Acta 1100: 15–26Google Scholar
  20. Hope AB, Matthews DB and Valente P (1994) The kinetics of reactions around the cytochromebf complex studied in an isolated system. Photosynth Res 40: 199–206Google Scholar
  21. Jeffrey GA and Saenger W (1991) Hydrogen Bonding in Biological Systems, p 159. Springer-Verlag New YorkGoogle Scholar
  22. Marcus RA and Sutin N (1985) Electron transfers in chemistry and biology. Biochim Biophys Acta 811: 265–322Google Scholar
  23. Martinez SE, Huang D, Cramer WA and Smith JL (1994) Crystal structure of cytochromef reveals a novel cytochrome fold and unexpected heme ligation. Structure 2: 95–105Google Scholar
  24. Niwa S, Ishikawa H, Nikai S and Takabe T (1980) Electron transfer reactions between cytochromef and plastocyanin fromBrassica komatsuna. J Biochem 88: 1177–1183Google Scholar
  25. Perutz MF (1974) Mechanism of denaturation of haemoglobin by alkali. Nature 247: 341–344Google Scholar
  26. Qin L and Kostic NM (1992) Electron-transfer reactions of cytochromef with flavin semiquinones and with plastocyanin. Importance of protein-protein electrostatic interactions and of donor-acceptor coupling. Biochemistry 31: 5145–5150Google Scholar
  27. Ratajczak R, Mitchell R and Haehnel W (1988) Properties of the oxidising site of Photosystem I. Biochim Biophys Acta 933: 306–318Google Scholar
  28. Rich PR (1982) Electron and proton transfers in chemical and biological quinone systems. Farad Disc Chem Soc 74: 349–364Google Scholar
  29. Rich PR (1985) Mechanisms of quinol oxidation in photosynthesis. Photosynth Res 6: 335–348Google Scholar
  30. Rich PR and Bendall DS (1980) The kinetics and thermodynamics of the reduction of cytochromec by substitutedp-benzoquinols in solution. Biochim Biophys Acta 592: 506–518Google Scholar
  31. Rich PR, Heathcote P and Moss DA (1987) Kinetic studies of electron transfer in a hybrid system constructed from the cytochromebf complex and Photosystem I. Biochim Biophys Acta 892: 138–151Google Scholar
  32. Rogers NK (1989) The role of electrostatic interactions in the structure of globular proteins. In: Fasman GD (ed) Prediction of Protein Structure and Principles of Protein Conformation. Plenum Press, New YorkGoogle Scholar
  33. Scheller HV and Møller BL (1990) Photosystem I polypeptides. Physiol Plant 78: 484–494Google Scholar
  34. Tamura N, Itoh S, Yamamoto Y and Nishimura M (1981) Electrostatic interaction between plastocyanin and P700 in the electron transfer reaction of photosystem I-enriched particles. Plant & Cell Physiol 22: 603–612Google Scholar
  35. Takabe T and Ishikawa H (1989) Kinetic studies of a cross-linked complex between plastocyanin and cytochromef. J Biochem 105: 98–102Google Scholar
  36. Takabe T, Niwa S, Ishikawa H and Tanaka Y (1980) Electron transfer reactions of cytochromef fromBrassica komatsuma with hexacyanoferrate. J Biochem 88: 1167–1176Google Scholar
  37. Takabe T, Ishikawa H, Niwa S and Tanaka Y (1984) Electron transfer reactions of chemically modified plastocyanin with P700 and cytochromef. Importance of local charge. J Biochem 96: 385–393Google Scholar
  38. Tikhonov AN, Khumotov GB, Ruuge EK and Blumenfeld LA (1981) Electron transport control in chloroplasts. Effects of photosynthetic control monitored by the intrathylakoid pH. Biochim Biophys Acta 637: 321–333Google Scholar
  39. Whitmarsh J, Bowyer JR and Crofts AR (1982) Modification of the apparent redox reaction between cytochromef and the Rieske iron-sulfur protein. Biochim Biophys Acta 682: 404–412Google Scholar
  40. Widger WR and Cramer WA (1991) The cytochromeb 6 f complex. In: Bogorad L and Vasil IK (eds) Cell Culture and Somatic Cell Genetics of Plants, vol 7B, The Molecular Biology of Plastids, 149–176. Acad Press, San DiegoGoogle Scholar

Copyright information

© Kluwer Academic Publishers. Printed in the Netherlands 1994

Authors and Affiliations

  • A. B. Hope
    • 1
  • P. Valente
    • 1
  • D. B. Matthews
    • 2
  1. 1.School of Biological SciencesFlinders UniversityAdelaideAustralia
  2. 2.School of Physical SciencesFlinders UniversityAdelaideAustralia

Personalised recommendations