Fluctuations of the proton-electromotive force across the inner mitochondrial membrane Joaquim Procopio Instituto de Cie ˆncias Biome ´dicas, Universidade de Sa ˜ o Paulo, Caixa Postal 66208, 05508-970, Sa ˜ o Paulo, SP, Brazil Jose ´ A. Forne ´ s * Instituto de Fı ´sica, Universidade Federal de Goia ´s, Caixa Postal 131, 74001-970, Goia ˆnia, GO, Brazil Received 17 October 1996 The intermembrane mitochondrial space IMMSis delimited by the inner and outer mitochondrial mem- branes and defines a region of molecular dimension where fluctuations of the number of free protons and of transmembrane voltage can give rise to fluctuations in the proton-electromotive force E PMF across the inner mitochondrial membrane IMM. We have applied the fluctuation-dissipation theorem to an electrical equiva- lent circuit consisting of a resistor R m in parallel with a capacitor C m representing the passive electrical properties of the IMM, in series with another capacitor C b representing the proton-buffering power of the IMMS fluid. An access resistance R a was defined as a link between the capacitor C b and the membrane. Average E PMF fluctuations across the IMM were calculated for different assumptions concerning the inter- membrane space dimensions. The calculated average E PMF fluctuations were in the vicinity of 100 mV for relaxation times in the few-microseconds range. The corresponding fluctuational protonic free energy is about 10 kJ/mole, which is comparable to the binding energy for protons in different transporters. This suggests that fluctuations in E PMF can be of relevance in the universe of forces influencing the molecular machinery embedded in the IMM. S1063-651X9701205-1 PACS numbers: 87.10.+e, 51.90.+r I. INTRODUCTION The synthesis of ATP in eukariotic cells occurs at the expense of proton flow driven by the proton electromotive force ( E PMF ) across ATP synthase molecules inserted in the inner mitochondrial membrane IMM. Proton gradients as high as 1 p H can occur at that location 1and superposition with electrical potential difference can lead to E PMF ’s attain- ing the 200-mV mark. Since the intermembrane space in most of its trajectory has a width of only about 10–20 nanometers, this dimension limits a region of molecular proportions where fluctuations in thermodynamic parameters may become an important part of the forces acting upon the molecular machines inserted alongside the inner mitochondrial membrane. This paper analyzes some consequences of the particular geometry of the intermembrane space IMSin mitochondria IMMS, which may result in relatively important fluctua- tions of proton-electromotive force across the IMM. II. THEORY The free-energy change G for the creation of an elec- trochemical gradient by an ion pump is 1the SI system of units is employed throughout G =RT ln c 2 c 1 +zF , 1 where c 2 c 1 -1 is the concentration ratio for the ion that moves, z is the ion valence, F is Faraday’s constant, R is the gas constant, T is the absolute temperature, and  is the transmembrane difference in electrical potential measured in volts. The proton-electromotive force is defined by E PMF =2.3 RT F p H+. 2 When G =0 zero chemical driving force, E PMF =0), Eq. 3can be used to relate the variations of p H across the membrane with voltage changes p H=- F  2.3RT =- e  2.3kT , 3 where e is the electronic charge and k the Boltzmann con- stant. In order to determine the fluctuations of the E PMF across the IMM, we need to know the susceptibility ( ) of the system comprised of the following elements see Fig. 1: 1 the inner mitochondrial membrane with its associated resis- tance and capacitance; 2the proton buffer compartment associated with the intermembrane fluid; 3the access resis- tance between the first two elements. In doing the above associations we can model the system as a set of electrical resistors and capacitors. The IMM and its surrounding solutions are represented by the membrane capacitor C m . The conductive pathways across the IMM, which include the pump channel and other leaks including decouplers, are collectively represented by the term R m . The buffering capacity of the intermembrane fluid is repre- sented by an electrical equivalent calculated belowdefined as C buffer =C b . This compartment is electrically connected to the IMM by an access resistance R a . *Corresponding author. PHYSICAL REVIEW E MAY 1997 VOLUME 55, NUMBER 5 55 1063-651X/97/555/62854/$10.00 6285 © 1997 The American Physical Society