Medium and Temperature Effects on the Redox Chemistry of Cytochromec

Abstract

Cytochromes c (cytc) are ubiquitous heme-containing metalloproteins that shuttle electrons in a variety of electron-transport chains, most often central to the production of the chemical energy necessary for cell life. The reduction potential (E°Ј) of the Fe 3+/2+ couple is central to the physiological role of these species in that it influences the thermodynamic and kinetic features of electron-exchange reactions with redox partners. In the last two decades, voltammetric techniques exploiting the heterogeneous electron exchange between cytc and solid electrodes have proved to be particularly valuable for the determination of E°Ј values for these species and for characterizing the mechanistic and kinetic aspects of the redox process for the various cytc conformers under a variety of solution conditions. The understanding of how, and to what extent, different molecular factors control the E°Ј value in these species has been the subject of much debate. First coordination sphere effects on the heme iron and the interactions of the heme group with the surrounding polypeptide [a] Gianantonio Battistuzzi obtained his Ph.D. in Chemistry from the University of Modena (Italy) in 1996, under the supervision of Prof. M. Sola, working on the purification and the characterization of the solution behavior of electron-transfer metalloproteins. After a period in the laboratories of Professors H. Witzel and B. Krebs at the University of Münster, he held a postdoctoral position at the University of Modena, where he was appointed Assistant Professor of Inorganic Chemistry in 1999. His main research interest lies in the understanding of the molecular and solution factors influencing the thermodynamics of the reduction of electron-transfer metalloproteins, in particular cytochromes c and blue copper proteins, through a combination of electrochemical and spectroscopic techniques. Marco Borsari obtained his Ph.D. in Chemistry from the University of Modena (Italy) in 1992, working with Prof. G. Gavioli on the characterization of the electrochemical behavior of sulfur-containing organic molecules and the determination of the nature and stability of metal complexes in solution through voltammetric and polarographic techniques. In 1992, he was appointed Assistant Professor of Physical Chemistry at the University of Modena and Reggio Emilia (Italy) and started working on the electrochemistry of metalloproteins with Marco Sola. He also established collaborations with the groups of I. Bertini and H. B. Gray. His main research activity has been directed towards the study of the redox properties of electron-transport proteins and redox enzymes, with a particular focus on the thermodynamics of the electron-transfer processes in cytochromes, blue copper, and ironϪsulfur proteins. Marco Sola obtained his Ph.D. in Chemistry from the University of Parma (Italy) in 1987. Between 1985 and 1991 he was a member of the group of I. Bertini and C. Luchinat in Florence working on NMR of paramagnetic metalloproteins. He spent 1988 as a research associate at ...

Redox thermodynamics of cytochromes c subjected to urea induced unfolding

“…This effect, in fact, could favour the loss of proton(s) responsible for the transition. For the X LT species, the free energy change for ferriheme reduction (DG 0 rc ) is determined by opposite enthalpic and entropic contributions (Tables 2, 3 and 4), as observed previously for the N form [4,33]. In this case, the magnitude of each contribution increases markedly with increasing urea concentration but these effects offset each other (compensatory behavior), therefore no relevant changes in DS 0 rc are observed.…”

“…It has been proposed that this thermal transition involves a conformer with a lower protonation number characterized by a reduced state stabilized on enthalpic bases. However, its E°0 value strongly decreases with increasing temperature due to a much higher entropy loss upon reduction [4,33,49]. A greater opening of the heme crevice in the oxidized state as compared to the low-T conformer and reorganizational effects which also include a change in orientation of the axial methionine ligand could be the structural responsible of the observed phenomenon [33,48].…”

“…Solvent reorganization effects, reduction-induced changes in the hydrogen bonding network within the hydration sphere of the cytochrome c and in protein flexibility are supposed to play the major role in determining the DS 0 rc values [4, 33, 36-38, 51, 52]. The negative DH 0 rc values, however, mostly arise from the stabilization of the ferroheme by ligand binding interactions and the hydrophobicity of the heme environment, but is also modulated by electrostatic interactions of the heme with protein surface and solution charges [4,33,[36][37][38][51][52][53]. The slight decrease in the E°0 values observed for the N ?…”

“…X LT transition are not available yet, direct interaction of urea molecules with the cytochrome c surface and substitution of water molecules at the protein-solvent interface probably play an important role in the compensative change of the thermodynamic parameters. The absence of perfect compensation is most likely related to the fact that the variations in reduction enthalpy depend also on changes in electrostatics at the metal site and the selective stabilization of the oxidized heme by the increasing solvent exposure [4,33,[36][37][38][51][52][53]. In the case of the N ?…”

“…The molecular determinants of the formal reduction potential E°0, along with effects of temperature, ionic strength, solvent composition, pH-induced conformational changes and ligand binding have been investigated extensively [4][5][6][7][8]. Significant insight into the functional properties of cytochrome c can also be gained from the investigation of the folding and unfolding process, in particular the mechanisms controlling the formation of the functionally active three-dimensional structure of the protein [9][10][11][12][13][14].…”