ABSTRACT
This work addresses the possible role of the cell membrane in the molecular mechanism of action of two salan-type ruthenium complexes that were previously shown to be active against human tumor cells, namely [Ru(III)(L1)(PPh3)Cl] and [Ru(III)(L2)(PPh3)Cl] (where L1 is 6,6'-(1R,2R)-cyclohexane-1,2-diylbis(azanediyl)bis(methylene)bis(3-methoxyphenol); and L2 is 2,2'-(1R,2R)-cyclohexane-1,2-diylbis(azanediyl)bis(methylene)bis(4-methoxyphenol)). One-component membrane models were first used, a disordered fluid bilayer of dioleoylphosphatodylcholine (DOPC), and an ordered rigid gel bilayer of dipalmitoylphosphatidylcholine. In addition, two quaternary mixtures of phosphatidylcholine, phosphatidylethanolamine, sphingomyelin and cholesterol were used to mimic the lipid composition either of mammalian plasma membrane (1:1:1:1 mol ratio) or of a cancer cell line membrane (36.2:23.6:6.8:33.4 mol ratio). The results show that both salan ligands L1 and L2 bind relatively strongly to DOPC bilayers, but without significantly affecting their structure. The ruthenium complexes have moderate affinity for DOPC. However, their impact on the membranes was notable, leading to a significant increase in the permeability of the lipid vesicles. None of the compounds compromised liposome integrity, as revealed by dynamic light scattering. Fluorescence spectroscopy studies revealed changes in the biophysical properties of all membrane models analyzed in the presence of the two complexes, which promoted an increased fluidity and water penetration into the lipid bilayer in the one-component systems. In the quaternary mixtures, one of the complexes had an analogous effect (increasing water penetration), whereas the other complex reorganized the liquid ordered and liquid disordered domains. Thus, small structural differences in the metal ligands may lead to different outcomes. To better understand the effect of these complexes in cancer cells, the membrane dipole potential was also measured. For both Ru complexes, an increase in the dipole potential was observed for the cancer cell membrane model, while no alteration was detected on the non-cancer plasma membrane model. Our results show that the action of the Ru(III) complexes tested involves changes in the biophysical properties of the plasma membrane, and that it also depends on membrane lipid composition, which is frequently altered in cancer cells when compared to their normal counterparts.
ABSTRACT
The monoheme c-type cytochrome PccH from Geobacter sulfurreducens, involved in the pathway of current-consumption in biofilms, was electrochemically characterized in detail. Cyclic voltammetry was used to determine the kinetics and thermodynamics properties of PccH redox behavior. Entropy, enthalpy and Gibbs free energy changes associated with the redox center transition between the ferric and the ferrous state were determined, indicating an enhanced solvent exposure. The midpoint redox potential is considerably low for a monoheme c-type cytochrome and the heterogeneous electron transfer constant rate reflects a high efficiency of electron transfer process in PccH. The midpoint redox potential dependence on the pH (redox-Bohr effect) was investigated, over the range of 2.5 to 9.1, and is described by the protonation/deprotonation events of two distinct centers in the vicinity of the heme group with pKa values of 2.7 (pKox1); 4.1 (pKred1) and 5.9 (pKox2); 6.4 (pKred2). Based on the inspection of PccH structure, these centers were assigned to heme propionic acids P13 and P17, respectively. The observed redox-Bohr effect indicates that PccH is able to thermodynamically couple electron and proton transfer in the G. sulfurreducens physiological pH range.
ABSTRACT
Geobacter bacteria have a remarkable respiratory versatility that includes the dissimilatory reduction of insoluble metal oxides in natural habitats and electron transfer to electrode surfaces from which electricity can be harvested. In both cases, electrons need to be exported from the cell interior to the exterior via a mechanism designated as extracellular electron transfer (EET). Several c-type cytochromes from G. sulfurreducens (Gs) were identified as key players in this process. Biochemical and biophysical data have been obtained for ten Gs cytochromes, including inner-membrane associated (MacA), periplasmic (PpcA, PpcB, PpcC, PpcD, PpcE and GSU1996) and outer membrane-associated (OmcF, OmcS and OmcZ). The redox properties of these cytochromes have been determined, except for PpcC and GSU1996. In this perspective, the reduction potentials of these two cytochromes were determined by potentiometric redox titrations followed by visible spectroscopy. The data obtained are taken together with those available for other key cytochromes to present a thorough overview of the current knowledge of Gs EET mechanisms and provide a possible rationalization for the existence of several multiheme cytochromes involved in the same respiratory pathways.
