Low Midpoint Potential Research Articles

Studies on energy and electron transfer systems in green photosynthetic bacterium Chloropseudomonas ethylica strain 2K. I. Isolation and characterization of cytochromes from Chloropseudomonas ethylica strain 2K.

Two cytochromes of the C-type, c-551.5 and c-555(550), were isolated from the soluble fraction of a green photosynthetic bacterium, Chloropseudomonas ethylica strain 2K, and highly purified by DEAE-cellulose and Amberlite CG-50 ion exchanger column chromatography. Cytochrome c-551.5 has absorption maxima at 551.5, 523 and 418nm in the reduced state and at 528, 408 and 351 nm in the oxidized state. This cytochrome is an auto-oxidizable, slightly basic protein having an isoelectric point of 7.77. It has a low mid-point redox potential of −209 mV at pH 7.0. The molecular weight of this protein is close to 11,500 and it contains two heme c groups per molecule of protein. These results indicate that cytochrome c-551.5 closely resembles cytochrome c8 from Desulfovibrio vulgaris (strain Hildenborough). Cytochrome c-555(550) has absorption maxima at 555, 523 and 417.5 nm in the reduced state and at 525, 412 and 358 nm in the oxidized state. This protein is very acidic, having an isoelectric point of 4.65. It has a mid-point redox potential of +103mV at pH 7.0. Its molecular weight is approximately 12,000. Cytochrome c-555(550) is similar to algal and higher plant f-type cytochromes except for redox potential. There is no evidence for the presence of a B-type cytochrome in Cps. ethylica strain 2K.

Chemical hypothesis for energy conservation in the mitochondrial respiratory chain II. redox properties of the coupling site enzymes at sites I and II

The chemical hypothesis for energy conservation in the mitochondrial membrane postulates three coupling sites as required by the ADP/0 stoichiometry observed in mitochondrial oxidative phosphorylation, and a fourth energy conservation at cytochrome a3 thermodynamically coupled specifically to sites II and III. The fourth site conserves the free energy of the reaction between oxygen and cytochrome a3 by binding the water molecules produced at coupling sites II and III during electron transport through the coupling site enzymes. These coupling site enzymes participate directly in the redox reactions of electron transport in the respiratory chain by means of the dithiol-disulfide redox couple. The free energy of electron transport is conserved in an oxidized form of the coupling site enzyme. Analysis of the thermodynamic relationships between the energized and de-energized forms of this enzyme shows that, under the postulates of this hypothesis, the energized form always has a midpoint potential more positive than the de-energized form. In general, the midpoint potential of the de-energized form is comparable to that of the carrier of lower midpoint potential at the coupling site, while the midpoint potential of the energized form is comparable to that of the carrier of higher midpoint potential. The chemical hypothesis thus predicts that an observed shift of midpoint potential of a coupling site enzyme to a more negative potential upon energization of the respiratory chain must be a case of reversed electron transport through the coupling site, and that an apparent midpoint potential is measured rather than the true one as defined by direct equilibration with the normal hydrogen electrode. The anhydride storage potential of the fully energized, oxidized coupling site enzyme, E s ∗∗, at sites II and III, derives from two sources: one is the midpoint potential span ΔE m between the electron transport carriers of the coupling site; the other is binding of the water molecule produced in the esterification reaction at the coupling site by the fourth site of energy conservation at cytochrome a3. There is a partially energized, oxidized form of the coupling site enzyme Es at sites II and III which conserves the free energy of electron transport through the site as − TΔS of a strained conformation of the protein. These two species form two redox couples E∗/E∗s, and E∗∗/ E SH ∗∗ with the same redox potential, since the free energy of water binding should be the same for the oxidized or reduced form of the coupling site enzyme. One path of energization, starting from the reduced, de-energized coupling site enzyme E SH is the following: E SH ⇌ E ∗ H→E S ∗→E S ∗∗ , the interconversion of ESH and E SH ∗ being essentially isergonic. The other path is: E SH ⇌ E SH ∗→ E SH ∗∗→E S ∗∗, in which abstraction of the water molecule occurs prior to oxidation. Existence of this pathway predicts that site II can be partially activated to E SH ∗∗ in oxygen pulse experiments by the rapid oxidation of cytochrome a 3, before any electron transport can take place through that site.