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. 2013 Oct 1;22(12):1775–1785. doi: 10.1002/pro.2377

Figure 6.

Figure 6

AlphaScreen luminescent competition binding assay with ALA-scan CGRP and AM peptides. A: Competition binding with Biotin-CGRP (50 nM) and MBP-RAMP1 ECD-(GS)5-CLR ECD-H6 (50 nM) in the presence of the indicated unlabeled ALA-scan peptides in the CGRP(23-37)NH2 backbone. B: As in Panel B, except for Biotin-AM (100 nM) and MBP-RAMP2 ECD-(GS)2-RS-(GS)2-CLR ECD-H6 (100 nM) in the presence of the indicated unlabeled ALA-scan peptides in the AM(37-52)NH2 backbone. Data shown are the average of duplicate samples. C: Amino acid sequence alignment of CGRP(23-37) and AM(37-52) summarizing the ALA-scan mutagenesis results. Resides that are identical between CGRP and AM are shown in bold and conservative substitutions are underlined. Residues where mutation to ALA most significantly diminished binding are highlighted with a * and residues where mutation to ALA diminished binding to a lesser, but still significant degree are highlighted with a ∧. The ? denotes that our finding of CGRP K35A diminishing binding is at odds with previous findings.