Modulation of ligand selectivity by mutation of the first extracellular loop of the human C5a receptor

Stuart A. Cain, Trent M. Woodruff, Stephen M. Taylor, David P. Fairlie, Sam Sanderson, Peter N. Monk

Research output: Contribution to journalArticle

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Abstract

The cyclic C5a receptor antagonist, phenylalanine [L-ornithine-proline-D-cyclohexylalanine-tryptophan-arginine] (F-[OPchaWR]), has 1000-fold less affinity for the C5a receptor (C5aR) on murine polymorphonuclear leukocytes than on human. Analysis of C5aR from different species shows that a possible cause of this difference is the variation in the sequence of the first extracellular loop of the receptor. The mouse receptor contains Y at a position analogous to P 103 in the human receptor, and D at G 105 . To test this hypothesis, we expressed human C5aR mutants (P 103 Y, G 105 D and the double mutant, P 103 Y/G 105 D) in RBL-2H3 cells and investigated the effects of these mutations on binding affinity and receptor activation. All three mutant receptors had a higher affinity for human C5a than the wild-type receptor, but showed no significant difference in the ability of F-[OPchaWR] to inhibit human C5a binding. However, all of the mutant receptors had substantially lower affinities for the weak agonist, C5a des Arg 74 (C5adR 74 ), and two altered receptors (G 105 D and P 103 Y/G 105 D) had much lower affinities for the C-terminal C5a agonist peptide analogue, L-tyrosine-serine-phenylalanine-lysine-proline-methionine-proline-leucine-D- alanine-arginine (YSFKPMPLaR). Although it is unlikely that differences at these residues are responsible for variations in the potency of F-[OPchaWR] across species, residues in the first extracellular loop are clearly involved in the recognition of both C5a and C5a agonists. The complex effects of mutating these residues on the affinity and response to C5a, C5adR 74 , and the peptide analogues provide evidence of different binding modes for these ligands on the C5aR.

Original languageEnglish (US)
Pages (from-to)1571-1579
Number of pages9
JournalBiochemical Pharmacology
Volume61
Issue number12
DOIs
StatePublished - Jun 15 2001

Fingerprint

Anaphylatoxin C5a Receptor
des-Arginine Complement C5a
Proline
Modulation
Ligands
Mutation
Ala-Leu-Pro-Met
Phenylalanine
Arginine
Ornithine
Leucine
Tryptophan
Alanine
Methionine
Serine
Lysine
Tyrosine
Neutrophils
Chemical activation
Peptides

Keywords

  • Antagonist
  • C5a
  • Complement
  • Mast cells
  • Receptor

ASJC Scopus subject areas

  • Biochemistry
  • Pharmacology

Cite this

Cain, S. A., Woodruff, T. M., Taylor, S. M., Fairlie, D. P., Sanderson, S., & Monk, P. N. (2001). Modulation of ligand selectivity by mutation of the first extracellular loop of the human C5a receptor. Biochemical Pharmacology, 61(12), 1571-1579. https://doi.org/10.1016/S0006-2952(01)00608-6

Modulation of ligand selectivity by mutation of the first extracellular loop of the human C5a receptor. / Cain, Stuart A.; Woodruff, Trent M.; Taylor, Stephen M.; Fairlie, David P.; Sanderson, Sam; Monk, Peter N.

In: Biochemical Pharmacology, Vol. 61, No. 12, 15.06.2001, p. 1571-1579.

Research output: Contribution to journalArticle

Cain, SA, Woodruff, TM, Taylor, SM, Fairlie, DP, Sanderson, S & Monk, PN 2001, 'Modulation of ligand selectivity by mutation of the first extracellular loop of the human C5a receptor', Biochemical Pharmacology, vol. 61, no. 12, pp. 1571-1579. https://doi.org/10.1016/S0006-2952(01)00608-6
Cain, Stuart A. ; Woodruff, Trent M. ; Taylor, Stephen M. ; Fairlie, David P. ; Sanderson, Sam ; Monk, Peter N. / Modulation of ligand selectivity by mutation of the first extracellular loop of the human C5a receptor. In: Biochemical Pharmacology. 2001 ; Vol. 61, No. 12. pp. 1571-1579.
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