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. 2019 Oct 15;8:e48339. doi: 10.7554/eLife.48339

Figure 3. PRYSPRY missense variants maintain antibody binding.

(A) Representative Isothermal titration calorimetry (ITC) trace of IgG Fc against WT TRIM21 PRYSPRY fitted to the one set of sites model. (B) Summary of PRYSPRY variant binding affinities to IgG Fc. Consistent with known binding mode, TRIM21 PRYSPRY binds IgG with a stoichiometry of 2:1 (Keeble et al., 2008). Data provided in Figure 3—source data 1.

Figure 3—source data 1.
elife-48339-fig3-data1.xlsx (433.6KB, xlsx)
DOI: 10.7554/eLife.48339.008

Figure 3.

Figure 3—figure supplement 1. Representative ITC traces of IgG Fc titration against each PRYSPRY variant.

Figure 3—figure supplement 1.

(A) N297H. (B) A390V. (C) G440R. (D) F446I. (E) K455E. (F) Q470K. (G) G471R. (H) Buffer control. (I) Binding signature plot. In most experiments, 40 µM of IgG Fc was titrated against 8 µM of PRYSPRY except for the F446I (D) where the concentrations were halved. Data provided in Figure 3—source data 1.

Figure 3—figure supplement 2. Purification of TRIM21 PRYSPRY protein.

Figure 3—figure supplement 2.

(A–E) Superdex 75 (HiLoad 16/60) size exclusion chromatograms for a selection of TRIM21 PRYSPRY variants. (A) WT. (B) N297H. (C) A390V. (D) G440R. (E) F446I. (F) Variant TRIM21 PRYSPRY proteins resolved by 4–12% SDS-PAGE developed in MES-SDS buffer and stained using InstantBlue.