SUBSECOND MODULATION OF FORMYL PEPTIDE-LINKED GUANINE-NUCLEOTIDE-BINDING PROTEINS BY GUANOSINE 5'-O-(3-THIO)TRIPHOSPHATE IN PERMEABILIZED NEUTROPHILS

Rapid activation of guanine nucleotide-binding protein (G protein)-med iated signal transduction mechanisms occurs in many tissues. The human neutrophil provides a useful model for studying the mechanisms of the se fast processes. Fluorescent chemotactic tetrapeptide and pentapepti de exhibit 30-50% quenching of fluorescence upon binding to the neutro phil formyl peptide receptor, and their binding affinity is strongly r egulated by the G protein G(i). We used rapid kinetic spectrofluoromet ric methods to study the assembly and disassembly of the ternary compl ex of ligand, receptor, and G protein in digitonin-permeabilized human neutrophils. Binding was studied up to 20 nm ligand, where the half-t ime for association was 1.2 sec. The rate constant of association was near that for diffusion-limited reactions of ligands and proteins, 2 x 10(7) m-1 sec-1. The rate of uncoupling of formyl peptide receptor fr om G protein in the presence of high concentrations of guanine nucleot ide was greater-than-or-equal-to 5 sec-1 (i.e., t1/2 of 0.14 sec). Thu s, disassembly of the formyl peptide receptor-G protein complex occurs in the millisecond time domain and may be faster than the next step i n the signal transduction process.