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. 2014 Feb 21;9(2):e89669. doi: 10.1371/journal.pone.0089669

Figure 9. Determination of the Ca2+ sensitivities of the interactions between BSGPERx and CaM.

Figure 9

(A & B) Typical experiment to determine EC50(Ca2+) value for BSGPERx-CaM interactions. Reactions contained 0.5 µM BSGPERx, 2 µM XRhod-5F, 0.25 mM BAPTA, and saturating CaM concentration (obtained from Kd titrations). Incremental aliquots of Ca2+ were added as the emission intensities of XRhod-5F (A, open circles) and biosensor ratios (B, closed circles) were simultaneously monitored. Fractional biosensor response were plotted against free Ca2+ calculated from XRhod-5F response as described under Experimental Procedure. C, Titrations showing Ca2+ sensitivity of the interactions between CaM and BSGPER83–93 (open triangles), BSGPER150–175 (open squares), BSGPER242–259 (closed circles), BSGPER330–351 (closed triangles). Fits were performed on aggregate data (shown) of at least three independent experiments. Starting reaction mix for BSGPER83–93 titrations contained 3 mM BAPTA instead of 0.25 mM BAPTA.