Extended Data Fig. 10: Proton leak into proteoliposomes monitored by lipid-linked pHrodo fluorophores.

Buffer was exchanged from pH 6 to pH 8 (outside) and a step-like approximately 50% decrease in fluorescence was observed, which we interpret as the quenching of lipid-linked pHrodo on the liposome exterior, followed by a slow decay in fluorescence (black squares). The data were fit with a single exponential function (red line) with a time constant of about 33 s, which we interpret as the slow leak of protons into the liposome. The gaps in the data are periods in the absence of illumination, to rule out photobleaching as a cause of the fluorescence decay we observed. These experiments confirm that the pH gradient was maintained during the full time-frame in which we measure transport. Experiments were performed three times with similar results.