Cyclic agonists and antagonists of C5A receptors and G Protein-coupled receptors
a technology of c5a receptors and g protein-coupled receptors, which is applied in the direction of cyclic peptide ingredients, animal/human proteins, mammal proteins, etc., can solve the problems of reducing the number of cyclic agonists and antagonists. , to achieve the effect of facilitating the assembly of a
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example 1
Structure-Activity Relationship of C5a Agonists
[0095]We have focused on the C-terminal residues of C5a, in order to explore structure-activity relationships in the search for peptide sequences with potent agonist activity. Many of these peptides are full agonists relative to C5a, but have markedly lower potency (Sanderson et al, 1994, 1995; Finch et al, 1997). Our initial structure-activity investigations have been particularly informative. Mutating the decapeptide C-terminus of C5a (SEQ ID NO: 1, C5a65-74, ISHKDMQLGR) twice with I65Y and H67F (eg. 2) led to enhancement of agonist potency by about 2 orders of magnitude. These results are summarised in Table 2. Analyses of Ramschandran plots and 2D NMR spectra for compound 2 suggested that certain structural features, namely a twisted “helix-like” backbone conformation for residues 65-69 and a β-turn for residues 71-74, might be responsible for activity. These preliminary results provided some insight to structural requirements for t...
example 2
NMR Structure of C5a Antagonist
[0100]We used two dimensional nuclear magnetic resonance spectroscopy to determine the three dimensional structure of 7 and found that while there is no discernible structure in water, there is evidence of a stable gamma-turn structure in dimethylsulfoxide.
[0101]The 1D 1H-NMR spectrum of peptide 7 in d6-DMSO at 24° C. shows 4 distinct resonances for amide-NH protons, as summarized in Table 3. To establish their possible involvement in intramolecular hydrogen bonds, a deuterium exchange experiment was performed by adding a 10-fold excess of D2O to the solution. Two of the amide-NH doublets disappeared immediately, along with resonances attributable to the N-terminal methylamine protons. However, the other two amide NH resonances, as well as a broad resonance at approximately 8.05 ppm, persisted for up to 6.5 hours (FIG. 2). These three slowly-exchanging protons are assigned to the amide NHs of Trp and d-Cha and the side chain amine of Lys, the slow exch...
example 3
Structure-Activity Relationships in Vitro
[0108]We also examined the receptor-binding and antagonist activity of the hexapeptide 7 for comparison with our compounds. The previous-report by Konteatis et al (1994) concerned the ability of 7 to compete with C5a binding to receptors on isolated PMN membranes (IC50 70 nM), which is not necessarily physiologically relevant. We examined competition between 7 and C5a using intact PMN cells, and found that, under these conditions, 7 binds with much lower receptor affinity of IC50 1.8 μM. We confirmed that 7 is a full antagonist with no agonist properties. These results are summarized in FIG. 5a and Table 4. The relative affinity (ratio) of 7 for the C5aR in intact PMNs in our assays was similar to that previously reported for isolated PMN membranes.
[0109]We have also found that 7 shows antagonist activity against both C5a (FIG. 5b) and a C-terminal agonist decapeptide analogue 4 (YSFKPMPLaR) (Finch et al, 1997) of the C-terminus C5a65-74, sug...
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