Mechanisms of electron transfer in two decaheme cytochromes from a metal-reducing bacterium (original) (raw)
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Electron tunneling properties of outer-membrane decaheme cytochromes from Shewanella oneidensis
Geochimica et Cosmochimica Acta, 2007
We have characterized the outer-membrane decaheme cytochromes OmcA and MtrC from Shewanella oneidensis MR-1 at the singlemolecule level using scanning tunneling microscopy (STM) and tunneling spectroscopy (TS). These cytochrome proteins are of great interest because they are thought to mediate bacterial electron transfer reactions in anoxic waters that control the reductive dissolution of oxide minerals. In our study, to characterize the electron transfer properties of these proteins on a model surface, the purified cytochromes were chemically immobilized as molecular monolayers on Au(111) substrates via a recombinant tetra-cysteine sequence as verified by X-ray photoelectron spectroscopy. Atomic force microscopy images confirm the monolayer films were $5-8 nm thick which is consistent with the apparent lateral dimensions of individual cytochrome molecules obtained with STM. Current-voltage TS of single cytochrome molecules revealed that OmcA and MtrC have different abilities to mediate tunneling current despite having otherwise very similar molecular and biochemical properties. These observations suggest that, based on their electron tunneling properties, the two cytochromes could have specific roles during bacterial metal reduction. Additionally, this study establishes single-molecule STM/TS as an effective means for revealing insights into biogeochemical redox processes in the environment.
The Journal of Physical Chemistry B, 2013
Nonexponential distance dependence of the apparent electron-transfer (ET) rate has been reported for a variety of redox proteins immobilized on biocompatible electrodes, thus posing a physicochemical challenge of possible physiological relevance. We have recently proposed that this behavior may arise not only from the structural and dynamical complexity of the redox proteins but also from their interplay with strong electric fields present in the experimental setups and in vivo (J. Am Chem. Soc. 2010, 132, 5769−5778). Therefore, protein dynamics are finely controlled by the energetics of both specific contacts and the interaction between the protein's dipole moment and the interfacial electric fields. In turn, protein dynamics may govern electron-transfer kinetics through reorientation from low to high donor−acceptor electronic coupling orientations. Here we present a combined computational and experimental study of WT cytochrome c and the surface mutant K87C adsorbed on electrodes coated with self-assembled monolayers (SAMs) of varying thickness (i.e., variable strength of the interfacial electric field). Replacement of the positively charged K87 by a neutral amino acid allowed us to disentangle protein dynamics and electron tunneling from the reaction kinetics and to rationalize the anomalous distance dependence in terms of (at least) two populations of distinct average electronic couplings. Thus, it was possible to recover the exponential distance dependence expected from ET theory. These results pave the way for gaining further insight into the parameters that control protein electron transfer.
Large scale domain movement in cytochrome bc 1: a new device for electron transfer in proteins
Trends in Biochemical Sciences, 2001
Cytochrome bc 1 of most bacteria and mitochondria, and the analogous cytochrome b 6 f of chloroplasts and cyanobacteria, are key components of respiratory and photosynthetic electron transport chains 1-3 . These evolutionarily conserved energy transducing enzymes, generally known as cytochrome bc complexes, transfer electrons (e − s) from a hydroquinone (QH 2 ) derivative (ubi-, mena-or plastohydroquinone) to a c-type cytochrome or plastocyanin, and contribute to the generation of an electrochemical proton [H + ] gradient, which subsequently drives ATP synthesis, and ion and metabolite transport. Unlike the succinate dehydrogenases and some hydroquinone oxidases, which contribute only to the formation of a pH gradient, cytochrome bc 1 also transfers electrons across the membrane, thereby generating an electrical membrane potential. Thus, this enzyme contributes both to the ∆pH and ∆Ψ components of the proton motive force (∆µH + ) 4 . The first idea about how cytochrome bc 1 generated ∆µH + was presented by Wikström and Berden almost 30 years ago 5 . This was followed by the breakthrough model of Peter Mitchell, called the protonmotive Q-cycle 6 . However, even today, the mechanism of action of the catalytic events within cytochrome bc 1 is not completely understood.
Interheme electron tunneling in cytochrome c oxidase
Proceedings of the National Academy of Sciences, 2010
Cytochrome c oxidase (CcO) is the terminal enzyme of the respiratory chain that catalyzes respiratory reduction of dioxygen (O 2 ) to water in all eukaryotes and many aerobic bacteria. CcO, and its homologs among the heme-copper oxidases, has an active site composed of an oxygen-binding heme and a copper center in the vicinity, plus another heme group that donates electrons to this site. In most oxidoreduction enzymes, electron transfer (eT) takes place by quantum-mechanical electron tunneling. Here we show by independent molecular dynamics and quantum-chemical methods that the heme-heme eT in CcO differs from the majority of cases in having an exceptionally low reorganization energy. We show that the rate of interheme eT in CcO may nevertheless be predicted by the Moser-Dutton equation if reinterpreted as the average of the eT rates between all individual atoms of the donor and acceptor weighed by the respective packing densities between them. We argue that this modification may be necessary at short donor/ acceptor distances comparable to the donor/acceptor radii.
Single-Molecule Mapping of Long-range Electron Transport for a Cytochrome b 562 Variant
Nano letters, 2011
Cytochrome b 562 was engineered to introduce a cysteine residue at a surface-exposed position to facilitate direct selfassembly on a Au(111) surface. The confined protein exhibited reversible and fast electron exchange with a gold substrate over a distance of 20 Å between the heme redox center and the gold surface, a clear indication that a long-range electron-transfer pathway is established. Electrochemical scanning tunneling microscopy was used to map electron transport features of the protein at the single-molecule level. Tunneling resonance was directly imaged and apparent molecular conductance was measured, which both show strong redox-gated effects. This study has addressed the first case of heme proteins and offered new perspectives in singlemolecule bioelectronics.
Journal of Bacteriology, 2006
To assess the structural association of two outer membrane-associated c-type decaheme cytochromes (i.e., OmcA [SO1779] and MtrC [SO1778]) and their ability to reduce soluble Fe(III)-nitrilotriacetic acid (NTA), we expressed these proteins with a C-terminal tag in wild-type S. oneidensis and a mutant deficient in these genes (i.e., ⌬omcA mtrC). Endogenous MtrC copurified with tagged OmcA in wild-type Shewanella, suggesting a direct association. To further evaluate their possible interaction, both proteins were purified to near homogeneity following the independent expression of OmcA and MtrC in the ⌬omcA mtrC mutant. Each purified cytochrome was confirmed to contain 10 hemes and exhibited Fe(III)-NTA reductase activity. To measure binding, MtrC was labeled with the multiuse affinity probe 4,5-bis(1,3,2-dithioarsolan-2-yl)fluorescein (1,2-ethanedithiol) 2 , which specifically associates with a tetracysteine motif engineered at the C terminus of MtrC. Upon titration with OmcA, there was a marked increase in fluorescence polarization indicating the formation of a high-affinity protein complex (K d < 500 nM) between MtrC and OmcA whose binding was sensitive to changes in ionic strength. Following association, the OmcA-MtrC complex was observed to have enhanced Fe(III)-NTA reductase specific activity relative to either protein alone, demonstrating that OmcA and MtrC can interact directly with each other to form a stable complex that is consistent with their role in the electron transport pathway of S. oneidensis MR-1.
Electron transfer in cytochrome c depends upon the structure of the intervening medium
Structure, 1994
Background: Long-distance electron-transfer (ET) reactions through proteins are involved in a great many biochemical processes; however, the way in which the protein structure influences the rates of these reactions is not well understood. We have therefore measured the rates of intramolecular ET from the ferroheme to a bis(2,2'-bipyridine)imidazoleruthenium(II) acceptor at histidine 39 or 54 in derivatives of yeast iso-l-cytochrome c, and studied the effect of an asparagine to isoleucine mutation at position 52, a residue situated between the heme and the electron acceptor. Results: The Fe 2 +-Ru 3 + rate constants demonstrate that residue 52 affects ET from the heme to His54 (Ile52 >Asn52), but not to His39 (Ile52 = Asn52). The enhanced Fe 2 +-Ru 3 + (His54) electronic coupling for the N52I/K54H protein is in good agreement with cr-tunneling calculations, which predict the length of the ET pathways between the heme and His54. Conclusion: The structure of the intervening medium between the heme and electron acceptors at the protein surface influences the donor-acceptor couplings in cytochrome c.
Electron transfer between redox active proteins and mineral oxides is important in a variety of natural as well as technological processes, including electron transfer from dissimilatory metal-reducing bacteria to minerals. One of the pathways that could trigger electron transfer between proteins and minerals is redox-linked conformation change. We present electrochemical evidence that mitochondrial cytochrome c (Mcc) undergoes significant conformation change upon interaction with hematite and indium-tin oxide (ITO) surfaces. The apparent adsorption-induced conformation change causes the protein to become more reducing, which makes it able to transfer electrons to the hematite conduction band. Although Mcc is not a protein thought to be involved in interaction with mineral surfaces, it shares (or can be conformed so as to share) some characteristics with multiheme outer-membrane cytochromes thought to be involved in the transfer of electrons from dissimilatory iron-reducing bacteria to ferric minerals during respiration. We present evidence that a 10.1 kDa monohoeme cytochrome isolated and purified from Acidiphilium cryptum, with properties similar to those of Mcc, also undergoes conformation change as a result of interaction with hematite surfaces.
Electron transfer by domain movement in cytochrome bc1
Nature, 1998
The cytochrome bc1 is one of the three major respiratory enzyme complexes residing in the inner mitochondrial membrane. Cytochrome bc1 transfers electrons from ubiquinol to cytochrome c and uses the energy thus released to form an electrochemical gradient across the inner membrane. Our X-ray crystal structures of the complex from chicken, cow and rabbit in both the presence and absence of inhibitors of quinone oxidation, reveal two different locations for the extrinsic domain of one component of the enzyme, an iron-sulphur protein. One location is close enough to the supposed quinol oxidation site to allow reduction of the Fe-S protein by ubiquinol. The other site is close enough to cytochrome c1 to allow oxidation of the Fe-S protein by the cytochrome. As neither location will allow both reactions to proceed at a suitable rate, the reaction mechanism must involve movement of the extrinsic domain of the Fe-S component in order to shuttle electrons from ubiquinol to cytochrome c1. Su...