Hm. Miettinen et al., THE LIGAND-BINDING SITE OF THE FORMYL PEPTIDE RECEPTOR MAPS IN THE TRANSMEMBRANE REGION, The Journal of immunology, 159(8), 1997, pp. 4045-4054
We propose that the N-formyl-Met-Leu-Phe binding site in the human neu
trophil formyl peptide receptor (FPR) lies in the predicted transmembr
ane region. We examined the expression, binding, and G protein couplin
g of 28 mutated forms of FPR in stably transfected Chinese hamster ova
ry cells. The amino acids we mutated are: 1) predicted to be oriented
toward the interhelical space; 2) analogous to those required for liga
nd binding in various other G protein-coupled receptors; 3) divergent
from lipoxin A, receptor, a low affinity receptor for formylated pepti
des; and 4) either highly conserved or divergent in other G protein-co
upled receptors. Some mutations resulted in intracellular retention, s
uggesting that the receptors were misfolded. Most mutated receptors th
at were transported to the plasmalemma bound f-Nle-Leu-Phe-Nle-Tyr-Lys
-fluorescein with affinities similar to the wild-type receptor (K-d =
6 nM). However, mutations L78A (helix II), D106N, L109A (helix III), T
157A (helix IV), R201A, 1204Y, and R205A (helix V), W254A and Y257A (h
elix VI), and F291A (helix VII) resulted in reduced affinities (K-d =
30-128 nM). Of these mutations, D106N, R201A, and R205A also appeared
to affect G protein coupling, suggesting that these residues may also
be involved in signal transduction and/or are essential for proper fol
ding of the molecule. Some of the FPR residues that appeared to be inv
olved in binding of formylated peptides were located at sites analogou
s to those identified in ligand binding to certain other G protein-cou
pled receptors. It is thus possible that several G protein-coupled rec
eptors have a common placement of ligand-binding amino acids.