2022/04/12 by Komlódi, Timea, Tretter, László
paper · doi:10.26124/mitofit:2022-0012
The protonmotive force pmF establishes the link between electrical and chemical components of energy transformation and coupling in oxidative phosphorylation in the mitochondrial electron transfer system. The electrical part is corresponding to the mitochondrial membrane potential ΔΨm and the chemical part is related to the transmembrane pH gradient ΔpH. Although the contribution of ΔpH to pmF is smaller than that of ΔΨm, ΔpH plays an important role in mitochondrial transport processes and regulation of reactive oxygen species production. Separate measurement of ΔΨm and ΔpH allows for calculation of pmF. Methods for monitoring ΔΨm such as fluorescence dyes are generally available, while determination of ΔpH is more challenging. In this review, we focus on the application of the fluorescence ratiometric method using the acetoxymethyl ester form of 2,7-biscarboxyethyl-5(6)-carboxyfluorescein (BCECF/AM) for real-time monitoring of the intramitochondrial pH in isolated mitochondria. Knowing the intra- and extramitochondrial pH allows for calculating the ΔpH. Application of specific ionophores such as nigericin or valinomycin, exerts the possibility to dissect the two components of the pmF in different directions. Furthermore, we tried to summarize those mitochondrial processes, such as production of reactive oxygen species, where the ΔpH has an important role.