Design and fine-tuning redox potentials of metalloproteins involved in electron transfer in bioenergetics

Abstract

Redox potentials are a major contributor in controlling the electron transfer (ET) rates and thus regulating the ET processes in the bioenergetics. To maximize the efficiency of the ET process, one needs to master the art of tuning the redox potential, especially in metalloproteins, as they represent major classes of ET proteins. In this review, we first describe the importance of tuning the redox potential of ET centers and its role in regulating the ET in bioenergetic processes including photosynthesis and respiration. The main focus of this review is to summarize recent work in designing the ET centers, namely cupredoxins, cytochromes, and iron-sulfur proteins, and examples in design of protein networks involved these ET centers. We then discuss the factors that affect redox potentials of these ET centers including metal ion, the ligands to metal center and interactions beyond the primary ligand, especially non-covalent secondary coordination sphere interactions. We provide examples of strategies to fine-tune the redox potential using both natural and unnatural amino acids and native and nonnative cofactors. Several case studies are used to illustrate recent successes in this area. Outlooks for future endeavors are also provided. This article is part of a Special Issue entitled Biodesign for Bioenergetics--the design and engineering of electronic transfer cofactors, proteins and protein networks, edited by Ronald L. Koder and J.L. Ross Anderson.

Keywords: Cupredoxin; Cytochromes; Iron–sulfur proteins; Metalloenzymes; Non-covalent interactions; Secondary coordination sphere.

Figure 1

Reduction potential range of metal centers in electron transfer metalloprotein. Adapted from ref. [10]

Figure 2

(a) Schematic representation of molecules involved in light stage of photosynthesis. (b) Z-scheme of photosynthesis showing the electron flow and the potential of redox pairs involved. Figures adapted with permission from Ref. [29] (Copyright © 2004, Rights Managed by Nature Publishing Group under license number 3636630566455) and [32] (Copyright © 1992, Kluwer Academic Publishers. Under license number 3636630756533), respectively.

Figure 3

(a) Schematic representation of molecules involved in aerobic electron transport chain. Figure adapted from Ref. [35] Copyright © 2012 Elsevier B.V. under license number 3644871014372 (b) Associated redox potentials.

Figure 4

(a) Design of a T1 Cu site in a de novo four helix bundle. Figure adapted with permission from Ref. [63] Copyright © 2010, American Chemical Society. (b) Redesign of blue T1 Cu site in Az to green and red T1 Cu site through mutation of axial Met. Figure reprinted with permission from Ref. [68] Copyright © 2010, American Chemical Society.

Figure 5

(a) X-ray structure of the designed Cu_A center in Az. (PDB ID: 1CC3). (b) De novo designed Cu_A center in a four helix bundle. Figure adapted wiwth permission from Ref. [77]. Copyright © 2012, American Chemical Society.

Figure 6

(a) Molecular model of a 4-helix bundle with two types of hemes (hemes a and b) bound to it. (Figure adapted with permission from Ref. [80] Copyright © 2000, American Chemical Society. (b) De novo designed globin fold. Blue, white and red colors show the hydrophilic, neutral, and hydrophobic residues in domain I of the globin fold. Adapted with permission from Ref. [89]. Copyright © 1999, American Chemical Society.

Figure 7

(a) X-ray structure of (a) redesigned pig myoglobin (PDB ID: 1MNI) and (b) redesigned glycophorin A, that bind to heme through bis-His axial ligation. Figures are adapted from Ref. [10]

Figure 8

Ball and stick model of different FeS clusters.

Figure 9

Sequence and structural model of a colied-coil helical structure with [4Fe-4S] binding site. The Cys residues are shown as red sticks in the structure and are highlighted in yellow in the sequence. Figure is reprinted with permission from Ref. [138] Copyright © 2009 Elsevier B.V. Under license number 3636621075094.

Figure 10

An electrochemical set up to provide control over ET between two systems, O₂ detection and lactose detection. Reprinted with permission from Ref. [140]. Copyright © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim under license number 3636620458132.

Figure 11

De novo design of amphiphilic helical bundles and their future application in proton or electron transfer. Adapted with permission from Ref. [141] Published by Elsevier Ltd. Under license 3636620217012.

Figure 12

(a) The interaction between cytochrome b₅ and Mb(+6) (reprinted with permission from Ref. [156] Copyright © 2010, American Association for the Advancement of Science under license number 3636620002029). (b) Representative models of binding of Mb and Mb(+6) to cytochrome b₅ (reprinted with permission from Ref. [157] Copyright © 2012, American Chemical Society).

Figure 13

(a) The arrangement of hemes in tetraheme c₅₅₄ with their corresponding redox potentials. (b) The redox potential windows of tetraheme c₅₅₄. Figures reprinted with permission from Ref. [176]. Copyright © 2013 Elsevier B.V. under license number 363611278854.

Figure 14

Effect of changing the proximal ligand in heme proteins. The more electron donating the proximal ligand is, the higher the redox potential will be.

Figure 15

Role of axial ligand in potential of cytochromes [194,195].

Figure 16

Changing the ruffling of the heme in H-NOX (heme nitric oxide-oxygen reductase) causes a change in the redox potential of the heme. Redox potential of the WT H-NOX is about 420 mV. (all values are reported vs. SHE). Figure is reprinted with permission from Ref. [204]. Copyright © 2010, American Chemical Society.

Figure 17

Structure of (a) Rieske center in complex bc₁ (PDB ID: 1BE3) and (b) a plant-type [2Fe-2S] ferredoxin (PDB ID: 3AV8). As shown in the figure, while the clusters look very similar, the Rieske center has replaced two of its thiolate ligands with imidazolate. Values for redox potential are obtained from Ref. [210] and [211], respectively.

Figure 18

(a) Overall structure of a cupredoxins (azurin, PDB ID: 4AZU). (b) Representative active site structure of cupredoxins. (c) Overall scheme of greek key β-barrel fold of cupredoxins (adapted from [117]). (d), (e) Representative UV-vis and EPR spectra of cupredoxins, respectively. (adapted from [21])

Figure 19

Role of hydrophobicity of the axial ligand in redox potential of cupredoxins. Moving from more hydrophobic to more hydrophilic (a to c), the redox potential will decrease [4,228,230]. Cu ion is shown as a purple ball, the primary ligands are shown in cyan and the axial ligand is shown in black stick and balls.

Figure 20

Comparison of the heme binding pocket of Mb and CcP. CcP has an Asp ligand that H-bonds to its proximal His and lowers the redox potential (potentials from Ref. [180] and [243], respectively)

Figure 21

Comparison of H-bonding network between a Rieske-type protein (naphthalene dioxygenase-NDO) and a Rieske protein (water soluble fragment of complex bc₁- ISF). The figure is reprinted with permission from Ref. [251].

Figure 22

The [4Fe-4S] cluster is more buried in HiPIPS (PDB ID: 1HRR ) vs. ferredoxins (PDB ID: 1DFD). One of the ligands in ferredoxin is completely exposed to water.

Figure 23

Unnatural Met derivatives that are incorporated into axial position of Az and their corresponding redox potentials. Figure is reprinted from Ref. [269].

Figure 24

Redox active derivatives of Tyr used by Lu and coworkers to study water oxidation in mimics of CcO in Mb.

Figure 25

Addition of electron withdrawing groups to heme results in increase in reduction potential of the heme and in O₂ reduction activity in a Mb model of CcO. Figure adapted with permission from Ref. [274] Copyright © 2014, American Chemical Society.

Figure 26

SCS mutants of azurin span a range of reduction potential never reported before. The effect of these mutations is additive. Adapted from [235]

Figure 27

(a) Contribution of different factors in determining the E° in azurin mutants. (b) Active vs. passive H-bond. Reprinted with permission from [290]. Copyright © 2012, American Chemical Society.

Figure 28

Plot of ET rate vs. driving force of intermolecular ET in several azurin mutants, Reprinted with permission from [292]. Copyright {2015} American Chemical Society.

Figure 29

Several residues in primary and secondary coordination sphere of Mb that were discussed in terms of their effects on E° (PDB ID: 1MBN)

Figure 30

Several residues in primary and secondary coordination sphere of CcP that were discussed in terms of their effects on E_m. (PDB ID: 2CYP). Residues in black are surface charged residues.

Figure 31

(a) [2Fe-2S] cluster in MitoNEET and its PCS and SCS ligands. (b) Reduction potetinal values of a series of MitoNEET mutants. * indicates extreme potentials. Adapted from ref. [303] copyright {2010} American Chemical Society.

Figure 32

(a) Lowering the E° value in Fe-SOD vs Mn-SOD with different metal ions. (b) Tuning the E° value of SOD by SCS mutations. Figures adapted from ref. [310] Copyright © 2008, American Chemical Society.