The different possible dispositions of the electron transfer components in electron transfer chains are discussed: random distribution of complexes and ubiquinone with diffusion-controlled collisions of ubiquinone with the complexes, random distribution as above, but with ubiquinone diffusion not rate-limiting, diffusion and collision of protein complexes carrying bound ubiquinone, and solid-state assembly. Discrimination among these possibilities requires knowledge of the mobility of the electron transfer chain components. The collisional frequency of ubiquinone-10 with the fluorescent probe 12-(9-anthroyl)stearate, investigated by fluorescence quenching, is 2.3 X 10(9) M-1 sec-1 corresponding to a diffusion coefficient in the range of 10(-6) cm2/sec (Fato, R., Battino, M., Degli Esposti, M., Parenti Castelli, G., and Lenaz, G., Biochemistry, 25, 3378-3390, 1986); the long-range diffusion of a short-chain polar Q derivative measured by fluorescence photobleaching recovery (FRAP) (Gupte, S., Wu, E. S., Höchli, L., Höchli, M., Jacobson, K., Sowers, A. E., and Hackenbrock, C. R., Proc. Natl. Acad. Sci. USA 81, 2606-2610, 1984) is 3 X 10(-9) cm2/sec. The discrepancy between these results is carefully scrutinized, and is mainly ascribed to the differences in diffusion ranges measured by the two techniques; it is proposed that short-range diffusion, measured by fluorescence quenching, is more meaningful for electron transfer than long-range diffusion measured by FRAP, or microcollisions, which are not sensed by either method. Calculation of the distances traveled by random walk of ubiquinone in the membrane allows a large excess of collisions per turnover of the respiratory chain. Moreover, the second-order rate constants of NADH-ubiquinone reductase and ubiquinol-cytochrome c reductase are at least three orders of magnitude lower than the second-order collisional constant calculated from the diffusion of ubiquinone. The activation energies of either the above activities or integrated electron transfer (NADH-cytochrome c reductase) are well above that for diffusion (found to be ca. 1 kcal/mol). Cholesterol incorporation in liposomes, increasing bilayer viscosity, lowers the diffusion coefficients of ubiquinone but not ubiquinol-cytochrome c reductase or succinate-cytochrome c reductase activities. The decrease of activity by ubiquinone dilution in the membrane is explained by its concentration falling below the Km of the partner enzymes. It is calculated that ubiquinone diffusion is not rate-limiting, favoring a random model of the respiratory chain organization.(ABSTRACT TRUNCATED AT 400 WORDS)