Peter Mitchell

The protonmotive Q cycle: A general formulation

A critical appraisal of the protonmotive Q* cycle [l] in the general context of the protonmotive function of cytochrome systems [2] has suggested that the Q cycle, as originally conceived [l] , lacked generality and was open to criticism because it was not yet adequately emancipated from its accidental origins. My object in this letter is to define the general principles of the protonmotive Q cycle more explicitly than before, thus facilitating either its experimental rejection or its further development and general application.

Protonmotive redox mechanism of the cytochrome b-c1 complex in the respiratory chain: Protonmotive ubiquinone cycle

There are two major interrelated difficulties that have hampered progress in our understanding and rational experimental exploration of the protonmotive and redox functions of the cytochrome b-cl region of the respiratory chain, using orthodox biochemical theory together with the conceptual facilities of the classical proton-translocating redox loop [l] and some variants of this [l-3] that have so far been explicitly at our disposal. One difficulty is, as pointed out by Slater [4] , the apparent lack of a hydrogen carrier for Loop 3. The other difficulty is the peculiar kinetic and thermodynamic behaviour of the b cytochromes, which has received much attention [S-l 11, but not yet a completely satisfactory explanation [ 12-211 . In this paper I describe the protonmotive Q* cycle that provides a basis, both for explaining the protontranslocating function of the cytochrome b-cl segment of the respiratory chain, and for rationalising much of the biochemical information about the behaviour and functions of the b cytochromes and ubiquinone, which previously appeared to be difficult to interpret simply.

A chemiosmotic molecular mechanism for proton-translocating adenosine triphosphatases.

Up to the present time, considerations of the possible mechanism of the reversible ATPases of mitochondria, chloroplasts and bacteria have generally depended, either on largely chemical arguments about the transitional intermediates of the reaction mechanism, or on largely thermodynamic arguments about the mechanism of reversal of the hydrolytic process [ 1171. In this paper, I have used both types of argument together, and have thus suggested a simple chemiosmotic molecular mechanism for the reversible proton-translocating ATPases. The proposed mechanism has several characteristics by which it may be recognised experimentally, andit may therefore help to stimulate further research on this subject.

Chemiosmotic Coupling in Energy Transduction: A Logical Development of Biochemical Knowledge

At the invitation of the Editors, this paper gives a summary sketch of my position regarding some metabolic aspects of energy transduction and describes some present and anticipated perspectives from my point of view. To maintain as broad a horizon as possible, however, I have used this opportunity to describe how my views, and the rationale that I have developed to express them, have been derived from accepted or acceptable physicochemical theory and biochemical knowledge stemming from the creative and painstaking observations of my progenitors and colleagues.

Performance and Conservation of Osmotic Work by Proton-Coupled Solute Porter Systems

According to the chemiosmotic coupling conception of oxidative and photosynthetic phosphorylation systems, 1–3 the hydrogen and electron carriers of the respiratory chain and photoredox chain are looped across the non-aqueous (proton-insulating) M phase of the coupling membrane in such a way that redox activity along the chain is accompanied by the translocation of protons from one side of the membrane to the other, generating a protonmotive force of some 200 to 300 mV between the aqueous (proton-conducting) phases on either side. Thus, the primary physiological function of the respiratory chain and photoredox chain systems of mitochondria, chloroplasts and microorganisms is regarded as the provision of a source of power in the form of the proton current that can be used by appropriate systems plugged through the M phase of the coupling membrane.4–7

Modulation of coupling factor ATPase activity in intact chloroplasts: The role of the thioredoxin system

Photophos~ho~lat~on has mostly been studied using washed, broken chloroplas~s which have lost their outer envelopes and stromal contents f I]_ These studies suggest that the enzyme which catalyses pho~ophosphorylation (coupling factor, or CF,--CF1) can exist in several states of differing activify depending on the ~~per~n~ental conditions used. In darkadapted chloroplasts, CF,-CF1 exists in a cat~lyti~fly inactive state, but is converted into an active state on impressing Ap(H*) across the thylakoid membrane, either in the form of ApH f’2,3] or as A

Proton-coupled β-galactoside translocation in non-metabolizingEscherichia coli

[4,5]. The presence of an H+-specific channel, or proton-well, through CFo effectively converts a A+ into a ApH across CF, [6]. Thus the active state of the anisotropic coupling factor complex may only be induced near a dual pH optimum, when the stromal side of CFr is close to pH8 and the CF, side is near pH 5, This situation corresponds to the normal environmental condition in illuminated intact ~hloroplas~s (i.e.? those that retain their outer envelopes and stromal contents [7]). In broken chloroplasts, the catalytic properties of CFo-CFr near the dual pH optimum depend on the

A commentary on alternative hypotheses of protonic coupling in the membrane systems catalysing oxidative and photosynthetic phosphorylation

Acid-base and electrogenic processes coupled to the flux of β-galactosides into non-metabolizing cells ofEscherichia coli have been studied.When β-glactoside was added to non-metabolizing suspensions ofE. coli, the pH of the suspension medium increased, indicating that the β-galactoside travelled in with acid equivalents. When the cells were made permeable to K+ ions, this inflow of acid equivalents was accompanied by an equal outflow of K+ ions, indicating that each acid equivalent carried one positive charge across the membrane, and corresponded to an H+ ion going in or an OH− ion coming out. The effective movement of H+ ions, caused either by a pH difference or by an electrical potential difference across the membrane of the cells, was specifically facilitated by the presence of β-galactoside. These effects of β-galactoside were abolished by N-ethyl maleimide, which is known to inhibit the specific β-galactoside translocation.The possible involvement of a Na+-β-galactoside symporter was ruled out by showing that the galactoside-induced inflow of acid was practically independent of Na+ ion concentration in the range 0.05–50.0 mM, and that Na+ ions did not flow into the bacteria under the influence of a β-galactoside concentration gradient.It is concluded that the β-galactoside translocation inE. coli is probably mediated by a β-galactoside-H+ symporter or by a β-galactoside/OH− antiporter.

Cytochrome c oxidase is not a proton pump.

The elucidation of the molecular mechanism of ADP phosphorylation by the reversible ATPase complexes during oxidative phosphorylation in bacteria, mitochondria and chloroplasts is one of the most interesting, and perhaps most difficult, problems of biochemistry. It therefore seems all the more important that the discussion of alternative feasible types of mechanism, which may stimulate and guide appropriate experimental research, should not be confused either by misunderstandings about the relevant conceptual models that have been proposed, or about the implications of such models when they are developed in sufficient detail to make them practically realistic and experimentally testable. Leaving aside purely mechanical or conformationally coupled models, two main alternative types of biochemical model have been described to account for the reversal of the ATPase reaction in the ATPase complexes. They are, on the one hand, the protonmotive chemiosmotic type of mechanism [l-3], which has been evolved in considerable detail from a membrane-dependent theory of transport [4-91; and on the other hand, the protonic anhydride mechanism of Williams [lo-131, which has been evolved in outline from a membrane-independent theory of ‘dislocated phases’ between neighbouring redox and reversible ATPase components in ‘chains of catalysts’ ([lo] and see [8,9] ). These alternative concepts involve a different approach to the chemical mechanism of ADP phosphorylation. They also involve a radically different view of the topological relationship between the reversible ATPase complexes and the redox complexes that act as the source of power for ADP phosphorylation. Recent arguments about the relative merits of the protonmotive chemiosmotic mechanism and Williams’ protonic anhydride mechanism have seemed to me to confuse as much as to resolve the major scientific issues with which we are concerned [14-221. I have therefore thought it appropriate in this paper to review the origins of the alternative concepts and the relationships between them, and attempt to examine briefly their relative compatibility with existing experimental knowledge.

Active/inactive state transitions of mitochondrial ATPase molecules influenced by Mg2+, anions and aurovertin.

We conclude that the reduction of O2 to 2 H2O by cytochrome c oxidase of rat liver mitochondria involves the translocation of 4-from cytochrome c at the outer surface of the cristae membrane per O2 reduced and protonated by 4 H+ ions that enter the reaction domain from the inner aqueous phase. This net electron-translocating function of cytochrome c oxidase plugged through the mitochondrial cristae membrane is not linked to a proton-pumping function, such as that proposed by Wikström [7,8].