Evolution of cytochromes and photosynthesis.

The following is an outline of the direction of research into the evolutionary origins of photosynthesis as revealed by the study of cytochromes c. Determination of the numbers of kinds of cytochromes, their structures, their functional roles, and their distribution are the principal kinds of data being collected and analyzed. A hypothesis on the origin of photosynthesis is presented.     

Dietary effects on cytochromes P450, xenobiotic metabolism, and toxicity.

The levels and activities of cytochrome P450 enzymes are influenced by a variety of factors, including the diet. In this article, the effects of selected non-nutritive dietary chemicals, macronutrients, micronutrients, and ethanol on cytochromes P450 and xenobiotic metabolism are reviewed in the light of our current understanding of the multiplicity and substrate specificity of cytochrome P450 enzymes. Although the mechanisms of action of several dietary chemicals on specific cytochrome P450 isozymes have been established, those for macro- and micronutrients are largely unknown. It is known, however, that specific nutrients may have varied effects on different cytochrome P450 forms and thus may affect the metabolism of various drugs differently. Nutritional deficiencies generally cause lowered rates of xenobiotic metabolism. In certain cases, such as thiamin deficiency and mild riboflavin deficiency, however, enhanced rates of metabolism of xenobiotics were observed. The effects of dietary modulation of xenobiotic metabolism on chemical toxicity and carcinogenicity are discussed.     

Bacterial 4-alpha-helical bundle cytochromes.

The biological functions of cytochrome c' and bacterioferritin, both haemoproteins with a common 4-alpha-helical bundle structure, are discussed and an example given of one of Kamen's laws, namely: comparative studies of prokaryotic cytochromes and their eukaryotic counterparts are useful. In the present case, the comparison is between bacterioferritin and its animal counterpart, haemoferritin.     

Structural homology of cytochromes c.

Cytochromes c from many eukaryotic and diverse prokaryotic organisms have been investigated and compared using high-resolution nuclear magnetic resonance spectroscopy. Resonances have been assigned to a large number of specific groups, mostly in the immediate environment of the heme. This information, together with sequence data, has allowed a comparison of the heme environment and protein conformation for these cytochromes. All mitochondrial cytochromes c are found to be very similar to the cytochromes c2 from Rhodospirillaceae. In the smaller bacterial cytochromes, Pseudomonas aeruginosa cytochrome c551 and Euglena gracilis cytochrome c552, the orientation of groups near the heme is very similar, but the folding of the polypeptide chain is different. The heme environment of these two proteins is similar to that of the larger bacterial and mitochondrial cytochromes. Two low-potential cytochromes, Desulfovibrio vulgaris cytochrome c553 and cytochrome c554 from a halotolerant micrococcus have heme environments which are not very similar to those of the other proteins reported here.     

The cytochromes of Acanthamoeba castellanii.

1. Low-temperature difference spectra of gradient-purified mitochondria of Acanthamoeba castellanii reveal the presence of cytochromes b-555, b-562 and c-549, with a-type cytochromes having a broad asymmetrical maximum at 602 nm; these components were also observed in specta of whole cells. 2. The a-type cytochromes are unusual in that they have split Soret absorption maxima (at 442 and 449 nm) and an uncharacteristic CO difference spectrum. 3. CO difference spectra of whole cells and 'microsomal' membranes show large amounts of cytochrome P-420 compared with cytochrome P-450. 4. Difference spectra in the presence of cyanide indicate the presence of an a-type cytochrome and two cyanide-reacting components, one of which may be cytochrome a3. 5. Whole-cell respiration in a N2/O2 (19:1) atmosphere was decreased by 50%, suggesting the presence of a low-affinity oxidase. This lowered respiration is inhibited by 50% by CO, and the inhibition is partially light-reversible; photochemical action spectra suggest that cytochrome a3 contributes to this release of inhibition. Other CO-reacting oxidases are also present. 6. The results are discussed with the view that cytochrome a3 is present in A. castellanii, but its identification in CO difference spectra is obscured by other component(s).     

Methionine-oxidized horse heart cytochromes c. I. Reaction with Chloramine-T,products, and their oxidoreduction properties

Spectroscopically, the modification of horse heart ferricytochrome c with N-chloro-4-toluolsul-fonamide (Chloramine-T, CT) occurs through a two-step process, the disruption of the methionine-80 sulfur-iron linkage and a reagent-independent change, an intramolecular rearrangement. Chromatographic purification of the preparation at a 2.5:1 reagent-to-protein ratio, pH 8.0–8.5, yields two major products, the F^II and F^III CT-cytochromes c. Both products contain modification of only the methionines, 80 and 65, to sulfoxides; both are monomeric, reduced by ascorbate, and the ferrous forms are oxidized by molecular oxygen and bind carbon monoxide. The redox potentials of F^II and F^III are 135 and 175±15 mV. The F^III is indistinguishable from the native protein in its binding and the electron donor property toward mammalian cytochrome c oxidase. It also binds nearly as effectively as the native protein to yeast cytochrome c peroxidase, but is a less efficient donor. It is, however, a poor electron acceptor from both mammalian cytochrome c reductase and chicken liver sulfite oxidase. F^II lacks cytochrome c oxidase activity and is also a poorer substrate for the other three enzymes. Both the derivatives are consistently better electron donors than acceptors. It is concluded that the binding of cytochrome c to cytochrome c oxidase and to cytochrome c peroxidase does not require the integrity of the methionine-80 sulfur linkage and that the complexation process has a finite degree of freedom with regard to the state of the heme crevice opening. The alterations of the oxidoreduction function have been analyzed in light of both prevailing models of cytochrome c function, the two-site model (one site for oxidizing and the other for reducing enzymes) and the single-site model (the same site for the oxidizing and reducing enzymes). These observations can be accommodated by either model, given the latitude that the binding domains for the oxidizing and the reducing enzymes have finite overlapping and nonoverlapping regions.     

Sequence variability in bacterial cytochromes c.

Cytochromes c are proteins that can be defined both phenotypically and by their possession of a characteristic sequence motif. Many sequences from bacterial sources are known, and new ones are being reported every year. An analysis can be made as to what fraction of new sequences are members of already known classes or subclasses, and how many map into previously uninhabited regions of sequence space.     

Liver microsomal cytochromes P-450 and azoreductase activity.

Hepatic microsomal azoreductase activity with amaranth (3-hydroxy-4[(4-sulfo-1-naphthalenyl)azo]-2,7-naphthalenedisulfonic acid trisodium salt) as a substrate is proportional to the levels of microsomal cytochrome P-450 from control or phenobarbital-pretreated rats and mice or cytochrome P-448 from 3-methylchol-anthrene-pretreated animals. In the "inducible" C57B/6J strain of mice, 3-methylcholanthrene and phenobarbital pretreatment cause an increase in cytochrome P-448 and P-450 levels, respectively, which is directly proportional to the increase of azoreductase activity. However, in the "noninducible" DBA/2J strain of mice, only phenobarbital treatment causes the increase both in cytochrome P-450 levels and azoreductase activity, while 3-methylcholanthrene has no effect. These experiments suggest that the P-450 type cytochromes are responsible for azoreductase activity in liver microsomes.     

Ligand binding properties of cytochromes c'.

The cytochromes c' bind CO, alkylisocyanides and CN- with rate and equilibrium constants which are 10(2)- to 10(6)-fold smaller than other high-spin hemoproteins. The decreased affinity for exogenous ligands is largely associated with steric interactions at the heme coordination site. While CO and alkylisocyanides bind noncooperatively to the dimeric Rhodospirillum molischianum cytochrome c', CO, alkylisocyanides and CN- appear to bind cooperatively to the dimeric Chromatium vinosum cytochrome c' due to a ligand-linked dimer-monomer dissociation equilibrium. The differences between the cytochromes c' are thought to be due to differences in amino acid residues near the heme coordination site and subunit interface.     

Biogenesis of respiratory cytochromes in bacteria.

Biogenesis of respiratory cytochromes is defined as consisting of the posttranslational processes that are necessary to assemble apoprotein, heme, and sometimes additional cofactors into mature enzyme complexes with electron transfer functions. Different biochemical reactions take place during maturation: (i) targeting of the apoprotein to or through the cytoplasmic membrane to its subcellular destination; (ii) proteolytic processing of precursor forms; (iii) assembly of subunits in the membrane and oligomerization; (iv) translocation and/or modification of heme and covalent or noncovalent binding to the protein moiety; (v) transport, processing, and incorporation of other cofactors; and (vi) folding and stabilization of the protein. These steps are discussed for the maturation of different oxidoreductase complexes, and they are arranged in a linear pathway to best account for experimental findings from studies concerning cytochrome biogenesis. The example of the best-studied case, i.e., maturation of cytochrome c, appears to consist of a pathway that requires at least nine specific genes and more general cellular functions such as protein secretion or the control of the redox state in the periplasm. Covalent attachment of heme appears to be enzyme catalyzed and takes place in the periplasm after translocation of the precursor through the membrane. The genetic characterization and the putative biochemical functions of cytochrome c-specific maturation proteins suggest that they may be organized in a membrane-bound maturase complex. Formation of the multisubunit cytochrome bc, complex and several terminal oxidases of the bo3, bd, aa3, and cbb3 types is discussed in detail, and models for linear maturation pathways are proposed wherever possible.     

Respiratory systems and cytochromes in Campylobacter fetus subsp. intestinalis.

Cell suspensions of Campylobacter fetus subsp. intestinalis grown microaerophilically in complex media consumed oxygen in the presence of formate, succinate, and DL-lactate, and membranes had the corresponding dehydrogenase activities. The cells and membranes also had ascorbate-N,N,N',N'-tetramethyl-p-phenylenediamine oxidase activity which was cyanide sensitive. The fumarate reductase activity in the membranes was inhibited by p-chloromercuriphenylsulfonate, and this enzyme was probably responsible for the succinate dehydrogenase activity. Cytochrome c was predominant in the membranes, and a major proportion of this pigment exhibited a carbon monoxide-binding spectrum. Approximately 60% of the total membrane cytochrome c, measured with dithionite as the reductant, was also reduced by ascorbate-N,N,N',N'-tetramethyl-p-phenylenediamine. A similar proportion of the membrane cytochrome c was reduced by succinate under anaerobic conditions, whereas formate reduced more than 90% of the total cytochrome under these conditions. 2-Heptyl-4-hydroxyquinoline-N-oxide inhibited reduction of cytochrome c with succinate, and the reduced spectrum of cytochrome b became evident. The inhibitor delayed reduction of cytochrome c with formate, but the final level of reduction was unaffected. We conclude that the respiratory chain includes low- and high-potential forms of cytochromes c and b; the carbon monoxide-binding form of cytochrome c might function as a terminal oxidase.     

Electrostatic and steric control of electron self-exchange in cytochromes c, c551, and b5.

The ionic strength dependence of the electron self-exchange rate constants of cytochromes c, c551, and b5 has been analyzed in terms of a monopole-dipole formalism (van Leeuwen, J.W. 1983. Biochim. Biophys. Acta. 743:408-421). The dipole moments of the reduced and oxidized forms of Ps. aeruginosa cytochrome c551 are 190 and 210 D, respectively (calculated from the crystal structure). The projections of these on the vector from the center of mass through the exposed heme edge are 120 and 150 D. For cytochrome b5, the dipole moments calculated from the crystal structure are 500 and 460 D for the reduced and oxidized protein; the projections of these dipole moments through the exposed heme edge are -330 and -280 D. A fit of the ionic strength dependence of the electron self-exchange rate constants gives -280 (reduced) and -250 (oxidized) D for the center of mass to heme edge vector. The self-exchange rate constants extrapolated to infinite ionic strength of cytochrome c, c551, and b5 are 5.1 x 10(5), 2 x 10(7), and 3.7 x 10(5) M-1 s-1, respectively. The extension of the monopole-dipole approach to other cytochrome-cytochrome electron transfer reactions is discussed. The control of electron transfer by the size and shape of the protein is investigated using a model which accounts for the distance of the heme from each of the surface atoms of the protein. These calculations indicate that the difference between the electrostatically corrected self-exchange rate constants of cytochromes c and c551 is due only in part to the different sizes and heme exposures of the two proteins.