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Abstract
Electron transfer (ET) between redox proteins is an essential process in the respiratory and photosynthetic transport chains. While intra-protein ET is well characterized, the experimental methods to investigate inter-protein ET are limited by the presence of the solvent and by the transient nature of the protein-protein interaction and ET event, which are averaged in protein ensembles. Wiring precisely oriented redox protein partners to the nanoscale electrodes of an electrochemical scanning tunneling microscope allows to record the time- and distance-dependence of the current flowing between them. These methods have revealed that the current flowing between individual protein pairs extends much beyond tunneling distances and that it is electrochemically gated. However, the mechanism of this process, and especially the identity of the charge carriers in the aqueous solution, remains to be elucidated. To determine the species involved in long distance charge transport between the redox partner proteins Cc and Cc1 of the respiratory chain, we have performed recordings as a function of pH, in heavy water solutions, and in degassed solutions. We observe that the spatial span and electrochemical gating of long-distance currents are reduced at high pH. Currents are also less extended in heavy water and at low oxygen concentration. Using the experimental structures of oxidized and reduced Cc and pKa calculations, we identify residues that can switch their availability to form hydrogen bonds with water and/or engage in proton-coupled electron transfer reactions. Overall, our results show that long distance currents between cytochrome c (Cc) and cytochrome c1 (Cc1) through the aqueous solution are assisted by superoxide anions and by protons, and we discuss possible molecular mechanisms and their relevance for interprotein ET.
Supplementary materials
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Supplementary Information
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Contents:
Figure S1. Diagrams of intraprotein vs intraprotein ET in the respiratory chain, ECSTM setup, and strategy for protein conjugation to the electrodes.
Figure S2. I-z curves and histograms at pH 6.6.
Figure S3. I-z curves and histograms at pH 7.4.
Figure S4. I-z curves and histograms at pH 8.0.
Figure S5. β values vs. EC gate potential.
Figure S6. I-z curves and histograms in D2O at pD 6.6.
Supplementary Table. Calculations of pKa (see spreadsheet online).
Figure S7. Protein sequence alignment of Cc proteins from bacteria to human.
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Title
Supplementary table 1 pKa table Cc red ox
Description
Supplementary Table. Calculations of pKa
Calculation of pKa values of cytochrome c in the reduced and oxidized forms using the computational tools PropKa3.5 and H++.
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