Extended Data Fig. 3: LIV-BP rapidly binds and responds to ligand.

a, Schematic of His-tagged LIV-BP^SS directly surface-immobilized. b, Representative fluorescence (donor in green and acceptor in red) and FRET (blue) traces from a single LIV-BP^SS sensor imaged at 15-ms time resolution during the rapid delivery (vertical dashed line) of 10 μM (subsaturating) leucine. c, Ensemble-average FRET efficiency (symbols), fit to a single exponential function with a time-constant of approximately 16 ms (red line). d, FRET values of an ensemble of surface-immobilized LIV-BP^SS molecules summed into a contour plot (scale at right), demonstrating rapid and uniform response to leucine addition. e, Schematic of His-tagged LIV-BP^WT directly surface-immobilized. f, Representative fluorescence (donor in green and acceptor in red) and FRET (blue) traces from a single LIV-BP^WT sensor imaged at 10-ms time resolution during the rapid delivery (vertical dashed line) of 3 μM (saturating) leucine. g, Ensemble-average FRET efficiency of leucine (grey squares), isoleucine (red circles) or valine (blue triangles). Data are fit to a single exponential function with a time-constant of approximately 27 ms for leucine (grey line), 25 ms for isoleucine (red line) and 32 ms for valine (blue line). h, FRET histogram of an ensemble of surface-immobilized LIV-BP^WT molecules responding to leucine summed into a contour plot (scale at right), demonstrating rapid and uniform response to leucine addition. i, Schematic of LIV-BP^WT encapsulated within the lumen of liposomes in identical conditions to those used for transport assays (Supplementary Methods). Liposomes were pre-incubated in 100 μg ml⁻¹ α-haemolysin for 15 min at room temperature. j, Encapsulated LIV-BP^WT FRET response to injection of 3 μM leucine, in which the time of mixing is marked (dashed grey line). The time constant of the observed FRET response (about 23 ms), fit as above (green line), was identical to LIV-BP^WT directly immobilized to the surface. All experiments were performed at least three times with similar results.