Targeted mutagenesis on PDGFRα-Fc identifies amino acid modifications that allow efficient inhibition of HCMV infection while abolishing PDGF sequestration
Fig 10
Combination of multiple mutations affecting PDGF binding reduces the affinity of PDGFRα-Fc for PDGF beyond detection.
Quantification of biochemical binding affinity of PDGFRα-Fc variants for PDGF-BB was assessed by microscale thermophoresis. Various concentrations of PDGFRα-Fc wildtype, PDGFRα-Fc V242K (A), PDGFRα-Fc I139E + V242K (B), PDGFRα-Fc Y206S + V242K (C) or PDGFRα-Fc I139E + Y206S + V242K (D) were mixed with 0.1 nmol/l (A) or 1 nmol/l (B to D) fluorescently labeled PDGF-BB. Binding curves were generated by analysis of the ratio (Δ Fnorm) of fluorescence at MST-on time (1.5–2.5) seconds over the steady-state fluorescence (F0) for each concentration. Measurements were performed at 60% excitation power (A) or 20% excitation power (B). Error bars indicate standard deviation from 3 replicate measurements.