Abstract
The binding of G protein to the N-formyl peptide receptor of human
neutrophils was investigated with site-specific synthetic peptides.
Peptide CT336(322) (322RALTEDSTQTSDTAT336) from the carboxyl-terminal
tail region of the receptor competed with the receptor for binding
to bovine Gi protein. The peptide competition was assayed by dissociation
of a GTP-sensitive, rapidly sedimenting (7S) form of receptor-G protein
complex as analyzed by velocity sedimentation on linear sucrose density
gradients. An IC50 of 590 microM was determined for CT336(322) peptide.
A control peptide, with the reverse sequence, rCT322(336) (336TATDSTQTSDETLAR322),
did not perturb the sedimentation of the reconstituted receptor-G
protein complex up to the highest tested concentration, 3 mM. Other
peptides tested, corresponding to central portions of the predicted
intracellular loop regions CII140(127) (127VLHPVWTQNHRTVS140) and
CIII239(227) (227KIHKQGLIKSSRP239) of the receptor, failed to dissociate
the reconstituted receptor-G protein complex. Control peptides from
the extracellular region EII184(170) (170KTGTVACTFNFSPWT184) and
an unrelated sequence matching a portion of neutrophil cytochrome
b, CYT306(296) (296KVVITKVVTHPFKTIE306), were also ineffective. Our
results suggest that the cytoplasmic tail of the formyl chemotactic
peptide receptor is involved in its coupling to the signal-transducing
G protein.
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