Fig. 4: Binding of rapamycin induces significant orthosteric and allosteric changes in the structural ensemble of FKBP-cpNanoLuc-FRB with coherent behavior.
From: Engineering and exploiting synthetic allostery of NanoLuc luciferase
A Fractional deuterium labelling at three mixing times (30, 300, and 3000 s) as a function of rapamycin concentration (0, 2, 4, 6, 8, 10, 15, and 20 µM) binding to 8 µM FKBP-cpNanoLuc-FRB. Protein domains shown below on the x-axis. Gly-rich linkers between domains as dashed vertical lines. B Unprocessed HDX-MS labelling data at 3000 s mixing time as a function of rapamycin concentration are shown for representative peptides of each cluster (except C6 rapamycin-independent regions). Fit to a Hill equation (black trace). C Rapamycin dose-response of each amino acid in the biosensor protein (as defined by EC50 and Hill coefficients from fit to Hill equation) was clustered with a k-means method. HDX-MS measurements were averaged per amino acid from peptide-level data and summed across all time points. Centroid of each cluster (C1–C7) denoted by x; Voronoi boundaries—blue lines. D Dendrogram of families from clustering in C. Y-axis—linkage (arbitrary units in Euclidean space). Source data are provided as a Source Data file.
