Chemical hypothesis for energy conservation in the mitochondrial respiratory chain

Abstract

A chemical hypothesis for energy conservation in the mitochondrial respiratory chain is presented with particular emphasis on coupling sites II and III located between cytochrome b and oxygen. The basic premise underlying this hypothesis is that electron transport from substrate to oxygen and the reaction which conserves free energy are coupled at three coupling sites, as required by the ADP/O stoichiometry observed with intact, phosphorylating mitochondria. The coupling sites themselves participate in the redox reactions of electron transport. While the stoichiometry of oxidative phosphorylation requires three coupling sites, the thermodynamics requires that the free energy available from the reaction of reduced cytochrome oxidase with oxygen be also conserved for this process. This hypothesis postulates that there is a fourth site of energy conservation at cytochrome oxidase which is thermodynamically coupled specifically to sites II and III. The coupling sites are postulated to be lipoprotein enzyme complexes, each specific to its region of the respiratory chain, which contain two tightly bound proteins, one which participates in the redox reaction, the other which acts as a transducer protein to store the free energy of electron transport. The redox protein has three thiol groups capable of forming two different disulfides when oxidized. One disulfide is a low energy form E S. Reduction of this primary disulfide to the trithiol form E SH releases one thiol to form a thiol ester with a neighboring carboxyl group with no net free energy change, but producing one molecule of water. The other two thiol groups are then oxidized to the second disulfide, an energy-rich protein whose highly strained confirmation is held in place by the disulfide crosslink. The free energy of electron transport is conserved as − TΔS in this high energy form E S ∗. The thiol ester group of E S ∗ contributes nothing to energy conservation, but acts as a functional group to facilitate energy transfer. Energy transfer takes place by transesterification of the thiol ester group of E S with a thiol and carboxyl group of the transducer protein E T to regenerate the primary disulfide Es and produce the thiol-ester crosslinked protein E ∼. It is at this stage that the water molecule is removed. Energy distribution can then take place by further transesterifications to produce enzymes whose energized state is characterized by an active ester crosslink. Mechanisms by which these mediate phosphorylation of ADP, reverse electron transport, and energy-linked transhydrogenation are presented. Conservation of the free energy available from the reaction of oxygen and reduced cytochrome oxidase is postulated to involve a conformational change of this protein to a state which binds with high affinity at specific sites the water molecules produced at coupling sites II and III by electron transport through the redox protein E s. The dehydrated form of the oxidized cytochrome oxidase is the high energy form, while the hydrated form is the low energy form. The reduced oxidase is assumed to have a low affinity for water, and the water molecules are released to the medium from the reduced form of this enzyme.