Redox biointeractome of cytochromes in respiration and photosynthesis

Abstract

Protein complex formation is at least a two-step process in which the formation of a final, well-defined complex entails the initial formation of a dynamic encounter complex. Highly transient complexes, with lifetimes in the order of microseconds-milliseconds, exhibit moderate or low binding affinities, with dissociation constants in the micromolar-millimolar range. Electron transfer reactions mediated by soluble redox proteins exchanging electrons between large membrane complexes in respiration and photosynthesis are excellent examples of transient interactions. Here, experimental approaches based on diamagnetic and paramagnetic Nuclear Magnetic Resonance (NMR) spectroscopy and/or Isothermal Titration Calorimetry, combined with computational methods, have been used to study the molecular recognition processes of particular redox complexes involved in respiration and photosynthesis. The studies presented in this PhD thesis go into detail about the structural and biophysical basis of the following redox complexes: cytochrome c¿cytochrome c1, cytochrome c-cytochrome c oxidase and cytochrome c-galactonolactone dehydrogenase interactions in respiration and the cytochrome c6¿cytochrome f adduct in photosynthesis. All these ET ensembles exhibit proper coupling between the redox centers although they differ in their dynamic behavior, which can be ascribed to its distinct functionality depending on the organism and its biological context. Moreover, post-translational regulation can alter the usual mechanisms of such proteins. In this work, the structure determination of a phosphomimetic mutant of cytochrome c has also been addressed by means of solution NMR. Needless to say, such a multidisciplinary methodology, combining experimental and computational methods, opens new perspectives in our understanding of the dynamic, transient adducts formed between proteins beyond the model systems herein analyzed. The current document has been prepared by following the guidelines required by the University of Seville to submit the PhD thesis as a collection of journal papers and book chapters. It consists of the following sections: II. The list of Abbreviations. III. The list of Publications and merits of the PhD candidate. IV. A brief Summary of the PhD thesis. V. An Introduction and the state-of-the-art. VI. A section covering the Objectives of the PhD thesis. VII. A brief description of the Results and Discussion, including main achievements and outcomes. VIII. The global Conclusions. IX. The list of References which have been cited in previous sections. X. The Appendix I, containing the published journal papers and an accepted book chapter, on which the PhD thesis is based. All papers were published in scientific journals indexed in Journal Citation Reports (JCR) database. XI. The Appendix II, containing an additional manuscript in preparation focused on the structure and dynamics of a phosphomimetic variant of cytochrome c. XII. The Appendix III, including three datasets deposited in the Biological Magnetic Resonance Data Bank (BMRB) and the Protein Data Bank (PDB).