Use of conjugates of bovine serum albumin with poly(alkylene oxide)s for solubilization of riboflavin ester.

Solubilization of 2',3',4',5'-tetrabenzoyl-5-acetyl-1,5-dihydroriboflavin (benzaflavin) by conjugates of BSA with poly(alkylene oxide)s [poly(ethylene glycol) and block co-polymers of ethylene oxide and propylene oxide (pluronics)] was investigated. Conjugates of BSA with pluronics were shown to have more solubilizing efficiency towards benzaflavin than BSA itself. Solubilized forms of benzaflavin are able to inhibit NADPH-dependent peroxidation of lipids in rat liver microsomes. A study of solubilized riboflavin ester transfer to mitochondria demonstrated that conjugates of BSA with pluronics may be advantageously employed for this purpose. Inhibitory properties of solubilized forms of benzaflavin were demonstrated by the study of their influence on the enzyme activity of D-amino-acid oxidase from pig kidney. The results show that solubilized forms of biologically active compounds based on conjugates of proteins with poly(alkylene oxide)s may be used for testing of chemical substances in biochemical systems.

The stoichiometry of proton translocation through H+-ATPase of rat-liver mitochondria.

The ratio of the number of protons transported directly by H+-ATPase of intact mitochondria and that of the hydrolyzed ATP (the proton translocation quotient) is determined. A special kinetic method which makes possible determination of ATPase and H+-translocase activities in the same experiment has been used to estimate the proton translocation quotient for mitochondrial H+-ATPase. The quotient is found to be, on average, equal to 3.31 mol/mol. It is shown that the protonophore 3,5-di-tert-butyl-4-hydroxybenzylidenemalononitrile can greatly decrease the proton translocation quotient. This supports our assumption concerning a labile coupling between H+-translocase and ATPase in mitochondria [I. A. Gorskaya, K. F. Sholtz, S. A. Moreva, A. V. Kotelnikova (1979) Biochemistry (Engl. Transl. Biokhimiya) 44, 765-770]. A decrease in the translocation quotient in the presence of the protonophore is likely to be due to the ability of this uncoupler to take back the translocated protons into the mitochondrial matrix before their release into the medium. An electrostatic model of the molecular mechanism of H+-translocase and ATPase coupling in the H+-ATPase complex is discussed. The model is in agreement with the results obtained: transport of more than two protons per each hydrolyzed ATP molecule, and variable efficiency of the process.