Interactomic affinity profiling by holdup assay: Acetylation and distal residues impact the PDZome-binding specificity of PTEN phosphatase
Fig 5
Conversion of the holdup binding intensities into affinities constants.
(A) The violin plot shows the distribution of all the back-calculated apparent peptide concentrations obtained when both a quantifiable and significant (>0.20) BI value by holdup and a dissociation constant by FP were available for a given PDZ/PBM pair. On each violin representation, the vertical line indicates the range of the distribution while the horizontal lines show the final mean peptide concentration and its final standard deviation after outlier exclusion (considering the 3σ rule). The final average peptide concentrations represented by the thick lines are used to convert the holdup BI values into KD. On the top are given the peptide concentrations, the numbers of data points with and without exclusion and the standard errors of the mean. (B) Comparison between the converted dissociation constants from the holdup assay and the dissociation constants directly measured by FP assay. The dotted line represents the perfect theoretical correlation. Since the data points are randomly distributed on both sides of this dotted line, the R2 is indicative of the goodness of fit.