Introduction:
Experimental evidence has shown that the rate of ATP synthesis correlates more strongly with proton movement along the surface of the inner mitochondrial membrane (IMM) than with the transmembrane proton concentration gradient in the bulk aqueous phase. This has prompted a revision of the classical chemiosmotic theory of mitochondrial ATP production. It has been proposed that lateral proton transport along the IMM surface generates an electric potential that drives ATP synthase activity. Moreover, recent studies have reported proton accumulation and storage on the membranes of the myelin sheath and endoplasmic reticulum (ER), suggesting that surface-based energy storage via absorbed protons may be a fundamental principle of cellular bioenergetics. In this study, we examined whether membranes composed of different types of phospholipids and those enriched with either acidic or basic bee venom proteins differ in their capacity to enhance proton absorption at the membrane surface.

Methods:
Five types of phospholipids commonly found in myelin sheath and ER membranes were purified using immunoaffinity chromatography. Anionic and basic protein isoforms were isolated from bee venom via CM Sephadex C-50 chromatography. Liposomal membranes were prepared through ultrasonic frequency dispersion of phospholipid suspensions. Proton absorption at the membrane surface was assessed by measuring pH differences between liposome suspensions and pure water.

Results:
Membranes of liposomes enriched with anionic proteins showed enhanced proton absorption compared to protein-free liposomal membranes. Among all tested conditions, phosphatidylethanolamine membranes enriched with basic proteins exhibited the highest proton absorption, while other basic protein-enriched membranes showed minimal absorption.

Conclusions:
These findings support a revised model of membrane-associated energy storage, which holds potential pharmacological implications. A deeper understanding of membrane-based energy accumulation mechanisms may facilitate the development of novel therapies targeting age-related and pathological impairments in cellular bioenergetics.