Walking the seven lines: binuclear copper A in cytochrome c oxidase and nitrous oxide reductase.

The enzymes nitrous oxide reductase (N2OR) and cytochrome c oxidase (COX) are constituents of important biological processes. N2OR is the terminal reductase in a respiratory chain converting N2O to N2 in denitrifying bacteria; COX is the terminal oxidase of the aerobic respiratory chain of certain bacteria and eukaryotic organisms transforming O2 to H2O accompanied by proton pumping. Different spectroscopies including magnetic resonance techniques, were applied to show that N2OR has a mixed-valent Cys-bridged [Cu1.5+(CyS)2Cu1.5+] copper site, and that such a binuclear center, called CuA, does also exist in COX. A sequence motif shared between the CuA center of N2OR and the subunit II of COX raises the issue of a putative evolutionary relationship of the two enzymes. The suggestion of a binuclear CuA in COX, with one unpaired electron delocalized between two equivalent Cu nuclei, was difficult to accept originally, even though regarded as a clever solution to many experimental observations. This minireview in honor of Helmut Sigel traces several of the critical steps forward in understanding the nature of CuA in N2OR and COX, and discusses its unique electronic features to some extent including the contributions made by the development of methodology and the discovery of a novel multi-copper enzyme. Left: X-band (9.130 GHz) and C-band (4.530 GHz, 1st harmonic display of experimental spectrum) EPR spectra of bovine heart cytochrome c oxidase, recorded at 20K. Right: Ribbon presentation of the CuA domain in cytochrome c oxidase and nitrous oxide reductase.

Ancient Ascaris DNA Sequences of Cytochrome B, Cytochrome C Oxidase Subunit 1, NADH Dehydrogenase Subunit 1, and Internal Transcribed Spacer 1 Genes from Korean Joseon Mummy Feces.

We analyzed Ascaris ancient DNA of cytochrome b, cytochrome c oxidase subunit 1, NADH dehydrogenase subunit 1, and internal transcribed spacer 1 genes extracted from the feces or precipitates of 15- to 18th-century Korean mummies. After multiple Ascaris genes in ancient samples were successfully amplified by polymerase chain reaction (PCR), consensus sequences could be determined by the alignment of the sequences of cloned PCR products. The obtained sequences of each gene were highly similar to those of Ascaris spp. reported thus far but were genetically distinct from Baylisascaris, Parascaris, and Toxascaris spp. The current report establishes that the genetic characteristics of the Ascaris spp. infecting pre-modern Korean societies were not uniform but were diverse to some degree.

A perspective on Peter Mitchell and the chemiosmotic theory.

In 1991 Peter Mitchell wrote a last article that summarised his views on the origin, development and current status of his chemiosmotic ideas. I here review some of his views of that time on structures and mechanisms of several key bioenergetic components in relation to the subsequent advances that have been made.

Cytochrome c oxidase: 25 years of the elusive proton pump.

Since its discovery [Nature 266 (1977) 271], the function of cytochrome c oxidase (and other haem-copper oxidases) as a redox-driven proton pump has been subject of both intense research and controversy, and is one of the key unsolved issues of bioenergetics and of biochemistry more generally. Despite the fact that the mechanism of proton translocation is not yet fully understood on the molecular level, many important details and principles have been learned. In the hope of accelerating progress, some of these will be reviewed here, together with a brief presentation of a novel proton pump mechanism, and of the emergence of a molecular basis for control of its efficiency.

The redox state of cytochrome oxidase in brain in vivo: an historical perspective.

Recent evidence suggests that cytochrome oxidase is partially reduced under resting conditions in the brain. Previous data, recorded over the past 30 years from intact brain using optical methods in the visible wavelength range, are consistent with this observation. These older data, while not conclusive in themselves, support the overall conclusions. The historical perspective on the experiments and controversies illustrates a number of useful principles. The first is that new methods tend to produce new observations that may be difficult to reproduce due to the uniqueness of the instrumentation. The second is that any new and different observations cannot be assimilated without an acceptable theoretical framework and, without assimilation can have little impact. Finally, the mechanisms which might explain why cytochrome oxidase may be more reduced than previously thought are still not fully developed and, therefore, the physiological significance of such reduction is not known.

Redox interactions in cytochrome c oxidase: from the "neoclassical" toward "modern" models.

Because of recent experimental data on the redox characteristics of cytochrome c oxidase and renewed interest in the role of cooperativity in energy coupling, the question of redox cooperativity in cytochrome c oxidase is reexamined. Extensive redox cooperativity between more than two redox centers, some of which are spectrally invisible, may be expected for this electron transfer coupled proton pump. Such cooperativity, however, cannot be revealed by the traditional potentiometric experiments based on a difference in absorbance between two wavelengths. Multiwavelength analyses utilizing singular value decomposition and second derivatives of absorbance vs. wavelength have revealed a stronger cooperativity than consistent with the "neoclassical" model, which allowed only for weak negative cooperativity between two equipotential one-electron centers. A thermodynamic analysis of redox cooperativity is developed, which includes the possibilities of strong cooperative redox interactions, the involvement of invisible redox centers, conformational changes, and monomer/dimer equilibrations. The experimental observation of an oxidation of one of the cytochromes (a3) with a decrease in applied redox potential is shown to require both strong negative cooperativity and the participation of more than two one-electron centers. A number of "modern" models are developed using the analytical approaches described in this paper. By testing with experimental data, some of these models are falsified, whereas some are retained with suggestions for further testing.