Extended Data Fig. 7: Mutation of filament interface residues.
From: Human PRPS1 filaments stabilize allosteric sites to regulate activity
a. Panel of negative stain EM sections of PRPS1 engineered mutations in phosphate buffer in the presence of the indicated ligands. b. Chromatography curves from a Superose 6 of PRPS1 and three engineered, filament-interface mutations. c. Assay performed in buffer containing: 50 mM Potassium HEPES pH 7.6, 6 mM MgCl2, 0.1 mM EDTA, 1 mM DTT, 0.1 mg/mL bovine serum albumin. Left: Activity assay of the three engineered mutations with or without 50 mM potassium phosphate, pH 7.6 (N = 4 technical replicates). Right: Ratio of 50 mM phosphate: 0 mM phosphate activities (V) from the panel to the left. d. Substrate kinetics of the three engineered mutations at protein concentrations with detectable catalytic activity. Assay performed in buffer containing: 50 mM Potassium Phosphate pH 7.6, 6 mM MgCl2, 0.1 mM EDTA, 1 mM DTT, 0.1 mg/mL bovine serum albumin. Triplicate readings of one well for a single replicate (N = 1 technical replicate) are shown as open circles. e. Kinetic parameters for the wild type protein and the three engineered mutations.
