Molecular bioenergetics deals with the construction, function and regulation of the powerhouses of life. The present overview sketches scenes and actors, farsighted goals and daring hypotheses, meticulous tool-making, painstaking benchwork, lucky discovery, serious scepticism, emphatic believing and strong characters with weak and others with hard arguments, told from a personal, admittedly limited, perspective. Bioenergetics will blossom further with the search focused on both where there is bright light for ever-finer detail and the obvious dark spots for surprise and discovery.

The mechanisms for Fe(III) oxide reduction by Geobacter species are of interest because Geobacter species have been shown to play an important role in Fe(III) oxide reduction in a diversity of environments in which Fe(III) reduction is a geochemically significant process. Geobacter species specifically express pili during growth on Fe(III) oxide compared with growth on soluble chelated Fe(III), and mutants that cannot produce pili are unable to effectively reduce Fe(III) oxide. The pili of Geobacter sulfurreducens are electrically conductive along their length under physiologically relevant conditions and exhibit a metallic-like conductivity similar to that observed previously in synthetic organic metals. Metallic-like conductivity in a biological protein filament is a previously unrecognized mechanism for electron transport that differs significantly from the more well-known biological strategy of electron hopping/tunnelling between closely spaced redox-active proteins. The multihaem c -type cytochrome OmcS is specifically associated with pili and is necessary for Fe(III) oxide reduction. However, multiple lines of evidence, including the metallic-like conductivity of the pili and the fact that OmcS molecules are spaced too far apart for electron hopping/tunnelling, indicate that OmcS is not responsible for long-range electron conduction along the pili. The role of OmcS may be to facilitate electron transfer from the pili to Fe(III) oxide. Long-range electron transport via pili with metallic-like conductivity is a paradigm shift that has important implications not only for Fe(III) oxide reduction, but also for interspecies electron exchange in syntrophic microbial communities as well as microbe–electrode interactions and the emerging field of bioelectronics.

The mechanism of electron transport in Geobacter sulfurreducens biofilms is a topic under intense study and debate. Although some proteins were found to be essential for current production, the specific role that each one plays in electron transport to the electrode remains to be elucidated and a consensus on the mechanism of electron transport has not been reached. In the present paper, to understand the state of the art in the topic, electron transport from inside of the cell to the electrode in Geobacter sulfurreducens biofilms is analysed, reviewing genetic studies, biofilm conductivity assays and electrochemical and spectro-electrochemical experiments. Furthermore, crucial data still required to achieve a deeper understanding are highlighted.

The free energy profile for electron flow through the bacterial decahaem cytochrome MtrF has been computed using thermodynamic integration and classical molecular dynamics. The extensive calculations on two versions of the structure help to validate the method and results, because differences in the profiles can be related to differences in the charged amino acids local to specific haem groups. First estimates of reorganization free energies λ yield a range consistent with expectations for partially solvent-exposed cofactors, and reveal an activation energy range surmountable for electron flow. Future work will aim at increasing the accuracy of λ with polarizable forcefield dynamics and quantum chemical energy gap calculations, as well as quantum chemical computation of electronic coupling matrix elements.

The AOX (alternative oxidase) is a non-protonmotive ubiquinol–oxygen oxidoreductase that couples the oxidation of ubiquinol with the complete reduction of water. Although it has long been recognized that it is ubiquitous among the plant kingdom, it has only recently become apparent that it is also widely found in other organisms including some human parasites. In this paper, we review experimental studies that have contributed to our current understanding of its structure, with particular reference to the catalytic site. Furthermore, we propose a model for the ubiquinol-binding site which identifies a hydrophobic pocket, between helices II and III, leading from a proposed membrane-binding domain to the catalytic domain.

Streptomyces spp. are known to produce various types of biologically active compounds including antibiotics, antiparasitic agents, herbicides and immunosuppressants. P450 (cytochrome P450) enzymes may have key roles in these biosynthetic and biotransformation reactions. Recent genomic analysis of Streptomyces coelicolor A3(2) indicates that S. coelicolor may have six ferredoxins (Fdxs), four putative Fdx reductases (FdRs) and 18 P450 genes. However, there are few clues to explain the mechanisms and functions of Streptomyces P450 systems. To solve these questions, we have expressed and purified five S. coelicolor P450s, four FdRs and six Fdxs in Escherichia coli . Of the purified P450s, CYP105D5 has fatty acid hydroxylation activity in a system reconstituted with putidaredoxin reductase and Fdx4 or with spinach FdR and spinach Fdx, although the reconstitutions with FdR2 or FdR3 and any of the Fdxs did not support CYP105D5-catalysed oleic acid hydroxylation. Elucidation of the detailed mechanisms of electron transport system for Streptomyces P450 may provide the perspective for usefulness of P450s as a biocatalyst.

Regulation of photosynthetic membrane synthesis in Rhodobacter sphaeroides 2.4.1 is dependent on the interactions of numerous regulatory elements, with two of the most important being the cbb 3 terminal oxidase and the PrrBAC two-component regulatory system. Here, we reveal that the cbb 3 terminal oxidase possesses extensive, additional regulatory activities under anaerobic conditions, and that the PrrBAC system is further involved in the regulation of the expression of more than 20% of the R. sphaeroides genome under anaerobic conditions, extending well beyond functions related to redox gene expression.

Shewanella spp. demonstrate great variability in the use of terminal electron acceptors in anaerobic respiration; these include nitrate, fumarate, DMSO, trimethylamine oxide, sulphur compounds and metal oxides. These pathways open up possible applications in bioremediation. The wide variety of respiratory substrates for Shewanella is correlated with the evolution of several multi-haem membrane-bound, periplasmic and outer-membrane c -type cytochromes. The 21 kDa c -type cytochrome CymA of the freshwater strain Shewanella oneidensis MR-1 has an N-terminal membrane anchor and a globular tetrahaem periplasmic domain. According to sequence alignments, CymA is a member of the NapC/NirT family. This family of redox proteins is responsible for electron transfer from the quinone pool to periplasmic and outer-membrane-bound reductases. Prior investigations have shown that the absence of CymA results in loss of the ability to respire with Fe(III), fumarate and nitrate, indicating that CymA is involved in electron transfer to several terminal reductases. Here we describe the expression, purification and characterization of a soluble, truncated CymA (‘CymA). Potentiometric studies suggest that there are two pairs of haems with potentials of -175 and -261 mV and that ‘CymA is an efficient electron donor for the soluble fumarate reductase, flavocytochrome c 3 .

Many proteins with one or more haem groups bound per polypeptide chain are called cytochromes. They function in electron transfer reactions and some are involved directly in the catalysis of chemical reactions, most prominently the reduction of oxygen to water in the terminal step of cell respiration. When unmodified haem is present the cytochromes are referred to as b -type, but if the haem is covalently attached to thiol groups of a Cys-Xaa-Xaa-Cys-His motif then the cytochrome is a c -type. Neither the purpose of this post-translational modification, nor the mechanisms of the machineries that are necessary for formation of the thioether bonds between protein and haem, are fully understood. In bacteria the c -type cytochromes function in the periplasm where they are involved in a range of electron transport activities, including the reactions of denitrification, in which nitrate is reduced sequentially via nitrite, nitric oxide and nitrous oxide to nitrogen gas. Other types of cytochromes have haem molecules with modifications to their porphyrin ring. These include the a-, d-, d 1 - and o -types. Although Keilin first described the a-, b- and c- types of cytochrome more than 60 years ago, we still do not have clear explanations as to why one type of haem moiety does not suffice for the requirements of mitochondrial, thylakoid and bacterial electron transport.