68] Electrometric measurement of rapid proton-transfer reactions in respiratory chains

Abstract

This chapter describes the methods for the rapid electrometric measurement of fast proton-transfer reactions in respiratory chains. The respiratory chains of mitochondria and bacteria are composed of hydrogen-transfer and electron-transfer redox systems. The study of the mechanism of redox reactions involving both an electron and a hydrogen carrier requires, as an adjunct to methods for following fast electron transfer processes, rapid and sensitive pH measurement. Measurement of small and rapid pH changes has gained further interest since the recognition that redox and hydro-anhydro energy-transducing systems can generate a pH difference across the membrane in which they are located—hence, the importance of understanding the mechanism and role of active proton translocation in biomembranes. Small pH changes can be measured with high sensitivity either electrometrically with glass electrodes or spectrophotometrically and fluorometrically with acid-base indicators. Dual-wavelength spectrophotometers in combination with light-flash generators and suitable acid-base indicators have been used for the measurement of rapid, small pH changes in photosynthetic systems.