The selectivity and structural determinants of peptide antagonists at the CGRP receptor of rat, L6 myocytes.
1. Potency orders were determined for a series of agonists and antagonists on the calcitonin gene-related peptide (CGRP) receptor of rat L6 myocytes. The agents tested were all shown to have been active against CGRP, amylin or adrenomedullin receptors. 2. AC187 had a pIC50 of 6.8 +/- 0.10, making it 14 fold less potent as an antagonist than CGRP8-37 (pIC50, 7.95 +/- 0.14). Amyline8-37 was equipotent to AC187 (pIC50, 6.6 +/- 0.16) and CGRP19-32 was 3 fold less potent than either (pIC50, 6.1 +/- 0.24). 3. [Ala11]-CGRP8-37 was 6 fold less potent than CGRP8-37, (pIC50, 7.13 +/- 0.14), whereas [Ala18]-CGRP8-37 was approximately equipotent to CGRP8-37 (pIC50, 7.52 +/- 0.15). However, [Ala11,Ala18]-CGRP8-37 was over 300 fold less potent than CGRP8-37 (pIC50, 5.30 +/- 0.04). 4. [Tyr0]-CGRP28-37, amylin19-37 and adrenomedullin22-52 were inactive as antagonists at concentrations of up to 1 microM. 5. Biotinyl-human alpha-CGRP was 150 fold less potent than human alpha-CGRP itself (EC50 values of 48 +/- 17 nM and 0.31 +/- 0.13 nM, respectively). At 1 microM, [Cys(acetomethoxy)2,7]-CGRP was inactive as an agonist. 6. These results confirm a role for Arg11 in maintaining the high affinity binding of CGRP8-37. Arg18 is of less direct significance for high affinity binding, but it may be important in maintaining the amphipathic nature of CGRP and its analogues.
Howitt
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Department of Pharmaceutical and Biological Sciences, Aston University, Aston Triangle, Birmingham.
Poyner
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DR
D000595:Amino Acid Sequence; D000818:Animals; D015740:Calcitonin Gene-Related Peptide / Q000031:analogs & derivatives* / Q000494:pharmacology*; D000077221:Calcitonin Gene-Related Peptide Receptor Antagonists; D002460:Cell Line; D000242:Cyclic AMP / Q000096:biosynthesis; D006801:Humans; D008565:Membrane Proteins / Q000819:agonists / Q000037:antagonists & inhibitors / Q000378:metabolism; D008969:Molecular Sequence Data; D018482:Muscle, Skeletal / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D010455:Peptides / Q000737:chemistry* / Q000494:pharmacology*; D051381:Rats; D058265:Receptors, Adrenomedullin; D018015:Receptors, Calcitonin Gene-Related Peptide / Q000819:agonists; D058287:Receptors, Islet Amyloid Polypeptide; D018000:Receptors, Peptide / Q000819:agonists / Q000037:antagonists & inhibitors / Q000378:metabolism; D013329:Structure-Activity Relationship
D000077221:Calcitonin Gene-Related Peptide Receptor Antagonists; D008565:Membrane Proteins; D010455:Peptides; D058265:Receptors, Adrenomedullin; D018015:Receptors, Calcitonin Gene-Related Peptide; D058287:Receptors, Islet Amyloid Polypeptide; D018000:Receptors, Peptide; D000242:Cyclic AMP; D015740:Calcitonin Gene-Related Peptide