Identification of an N-formyl peptide receptor ligand binding domain by a gain-of-function approach.

Autor: Quehenberger O; Department of Immunology, The Scripps Research Institute, La Jolla, California 92037, USA., Pan ZK, Prossnitz ER, Cavanagh SL, Cochrane CG, Ye RD
Jazyk: angličtina
Zdroj: Biochemical and biophysical research communications [Biochem Biophys Res Commun] 1997 Sep 18; Vol. 238 (2), pp. 377-81.
DOI: 10.1006/bbrc.1997.7298
Abstrakt: Replacement of N-formyl peptide receptor (FPR) domains with those from a homologous receptor, FPR2, resulted in chimeric receptors displaying low binding affinity to fMet-Leu-Phe (fMLF). To characterize fMLF binding domain, we adopted a "gain-of-function" approach by selective replacement of non-conserved residues in the FPR2 portion of the chimeric receptors with those from the FPR. This led to the identification of 3 clusters of residues required for high-affinity fMLF binding. Introduction of 2 positively charged amino acids, Arg84 and Lys85, dramatically improved binding affinity of one chimeric receptor (Kd from 105 nM to 1.6 nM). Similarly, restoration of either Gly89/His90 or Phe102/Thr103 improved the binding affinity of another chimeric receptor from a Kd of 275 nM to a 2.3 Kd and 3.3 nM, respectively. Increased ligand binding affinity was accompanied by a gain in calcium mobilization capability, suggesting functional coupling to G proteins. These results demonstrate the presence of structural determinants in the first extracellular loop and its adjacent transmembrane domains that are essential for high affinity fMLF binding.
(Copyright 1997 Academic Press.)
Databáze: MEDLINE
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