Identification of the key residue of calcitonin gene related peptide (CGRP) 27-37 to obtain antagonists with picomolar affinity at the CGRP receptor.
Calcitonin gene related peptide (CGRP) plays an important role in the CNS and in the cardiovascular system. To identify high-affinity antagonists in competitive binding studies, we identified a novel radioactive tracer, [(3)H-propionyl-K(24)]-halphaCGRP 8-37, which was labeled in solution by a recently developed strategy using photolabile protecting groups at reactive side chains. This tracer was shown to be as potent as commercially available (125)I-tracers for the determination of agonists and to have increased sensitivity for antagonists. We applied it to investigate the predicted turn structures centered at Pro(29) and Pro(34). The substitution at positions 29 and 34 by turn-inducing amino acid mimetica showed that these turns are highly diverse. At position 29, a hydrophobic residue is preferred that constricts the secondary structure, whereas position 34 is required to stabilize the conformation of the backbone. All high-affinity analogues showed antagonistic properties with potency similar to CGRP 8-37.
Lang
Manja
M
Institute of Biochemistry, University of Leipzig, D-04103 Leipzig, Germany.
De Pol
Silvia
S
Baldauf
Carsten
C
Hofmann
Hans-Jörg
HJ
Reiser
Oliver
O
Beck-Sickinger
Annette G
AG
D000595:Amino Acid Sequence; D001667:Binding, Competitive; D015740:Calcitonin Gene-Related Peptide / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology; D000077221:Calcitonin Gene-Related Peptide Receptor Antagonists; D006801:Humans; D007553:Isotope Labeling; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology; D017433:Protein Structure, Secondary; D011869:Radioligand Assay; D018015:Receptors, Calcitonin Gene-Related Peptide / Q000819:agonists; D013329:Structure-Activity Relationship; D014316:Tritium
D000077221:Calcitonin Gene-Related Peptide Receptor Antagonists; D010446:Peptide Fragments; D018015:Receptors, Calcitonin Gene-Related Peptide; C446021:calcitonin gene-related peptide (27-37):; C508797:calcitonin gene-related peptide (8-37), propionyllysyl(24)-; D014316:Tritium; C058636:calcitonin gene-related peptide (8-37); D015740:Calcitonin Gene-Related Peptide