Pump proteins rely on the input, transmission, and dissipation of energy to carry out directional transport of ions across a membrane. In this work, we use multidimensional single-molecule spectroscopy to quantify the nonequilibrium thermodynamics for portions of the reaction cycle of bacteriorhodopsin. We demonstrate the role of microscopic irreversibility in the mechanism of action for optically driven pumps.
Biological machinery relies on nonequilibrium dynamics to maintain stable directional fluxes through complex reaction cycles. For such reaction cycles, the presence of microscopically irreversible conformational transitions of the protein, and the accompanying entropy production, is of central interest. In this work, we use multidimensional single-molecule fluorescence lifetime correlation spectroscopy to measure the forward and reverse conformational transitions of bacteriorhodopsin during trans-membrane H+ pumping. We quantify the flux, affinity, enthalpy and entropy production through portions of the reaction cycle as a function of temperature. We find that affinity of irreversible conformational transitions decreases with increasing temperature, resulting in diminishing flux and entropy production. We show that the temperature dependence of the transition affinity is well fit by the Gibbs-Helmholtz relation, allowing the ΔHtrans to be experimentally extracted.Zum Volltext