How Cytochromes with Different Folds Control Heme Redox Potentials (original) (raw)

The electrochemical midpoint potentials (E m 's) of 13 cytochromes, in globin (c, c 2 , c 551 , c 553), four-helix bundle (c′, b 562), R roll (b 5), and sandwich (f) motifs, with E m 's spanning 450 mV were calculated with multiconformation continuum electrostatics (MCCE). MCCE calculates changes in oxidation free energy when a heme-axial ligand complex is moved from water into protein. Calculated and experimental E m 's are in good agreement for cytochromes with His-Met and bis-His ligated hemes, where microperoxidases provide reference E m 's. In all cytochromes, E m 's are raised by 130-260 mV relative to solvated hemes by the loss of reaction field (solvation) energy. However, there is no correlation between E m and heme surface exposure. Backbone amide dipoles in loops or helix termini near the axial ligands raise E m 's, but amides in helix bundles contribute little. Heme propionates lower E m 's. If the propionic acids are partially protonated in the reduced cytochrome, protons are released on heme oxidation, contributing to the pH dependence of the E m. In all cytochromes studied except b 5 's and low potential globins, buried side chains raise E m 's. MCCE samples ionizable group protonation states, heme redox states, and side chain rotamers simultaneously. Globins show the largest structural changes on heme oxidation and four-helix bundles the least. Given the calculated protein-induced E m shift and measured cytochrome E m the five-coordinate, His heme in c′ is predicted to have a solution E m between that of isolated bis-His and His-Met hemes, while the reference E m for His-Ntr ligands in cytochrome f should be near that of His-Met hemes. Cytochromes are a diverse family of heme-containing proteins that are components of electron transfer chains (1, 2). There are soluble, extrinsic, and intrinsic membrane cytochromes. While they are usually all R (3), they are also found in R and all folds. Cytochrome heme electrochemical midpoint potentials (E m 's) 1 vary between-400 and +400 mV (vs SHE) (4-6). This 800 mV E m span represents an 18.8 kcal/mol change in free energy of heme ionization, equivalent to changing an active site residue pK a by 13.8 pH units. The range of cytochrome redox potentials results from differences in stabilization of the buried, cationic, oxidized heme by proteins in different motifs (7, 8). Redox reactions of cofactors such as hemes, quinones (9-11) and iron-sulfur complexes (12-15) have been studied by continuum electrostatics (16), free energy perturbation and microscopic LRA (17, 18), molecular dynamics (19, 20), quantum mechanical (14, 21), and QM/MM (22) techniques. Each cofactor type and analysis method provides different insights into the role of the local ligand geometry (15), backbone dipoles (12, 13, 23), side chains (14), and protein † Supported by the National Science Foundation MCB 0212696. K.H. acknowledges a Feodor Lynen Fellowship from the Alexander von Humboldt Foundation.