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. Author manuscript; available in PMC: 2014 Sep 20.
Published in final edited form as: ACS Chem Biol. 2013 Jun 26;8(9):1955–1963. doi: 10.1021/cb400274z

Figure 3. Sulfation of Tyr21 improves sulfopeptide binding affinity and modulates full-length receptor activity.

Figure 3

(A) The largest chemical shift perturbations (orange) are consistent with the putative sTyr21 sulfopeptide (green) binding site on CXCL122 (PDB ID 2K05). (B) Chemical shift perturbations induced by sulfated and unsulfated peptides were fitted to a quadratic binding equation to yield Kd values. The sTyr21 sulfopeptide (circles) bound CXCL12WT with Kd = 1.8 ± 0.2 mM (black), CXCL121 with Kd = 1.6 ± 0.2 mM (blue) and CXCL122 with Kd = 211 ± 23 µM (red). The Tyr21 peptide (triangles) bound CXCL12WT with Kd = 2.7 ± 0.5 mM (black), CXCL121 with Kd = 1.5 ± 0.4 mM (blue) and CXCL122 with Kd = 831 ± 137 µM (red). (C) The calcium response of FLAG-tagged CXCR4 variants was measured as a function of CXCL12WT concentration. Data are representative of two experiments each performed with three replicates. (D) Four parameter fits yielded each CXCR4 variants EC50 and maximum calcium response. CXCR4 variants with EC50 or maximum calcium response values more than three standard deviations from the mean CXCR4WT quantities are indicated with an asterisk.