Guanine nucleotides modulate the binding affinity of the oligopeptide chemoattractant receptor on human polymorphonuclear leukocytes.

Abstract

A B S T R A C T The oligopeptide chemoattractant receptor on human polymorphonuclear leukocyte (PMN) membranes exists in two affinity states. Since guanine nucleotides regulate the binding affinity and transductional activity of several other types of receptors, we examined the effect of nucleotides on the binding of N-formyl-methionyl peptides to their receptors on human PMN membranes. The addition of guanylylimidodiphosphate (0.1 mM), a nonhydrolyzable derivative of guanosine triphosphate (GTP), to PMN membrane preparations reduced the fraction of high-affinity receptors detected in equilibrium binding studies from 21.3±0.13 to 11.8±0.05% (P < 0.03), without altering the binding affinities. Since the total number of receptors remained unchanged, the effect of guanylylimidodiphosphate was to convert a portion of the receptors from the high-affinity state to the low-affinity state. At the maximal concentration of guanine nucleotide tested, '50% of the high-affinity sites were converted to low-affinity sites. The findings obtained by equilibrium binding were supported by kinetic studies since the dissociation of the radiolabeled oligopeptide chemoattractant N-formyl-methionyl-leucyl-[3H]phenylalanine from PMN membranes was accelerated in the presence of guanine nucleotide. The effect of guanine nucleotides was reversed upon washing, indicating that affinity conversion is bidirectional. The guanine nucleotide effects were greatest with nonhydrolyzable derivatives of GTP followed by GTP then guanosine diphosphate. Neither guanosine monophosphate nor any adenine nucleotide tested had an Address reprint requests to Dr. R. Snyderman.

Dual Mg2+ control of formyl‐peptide‐receptor–G‐protein interaction in HL 60 cells

“…This effect is due to a reduction in the number of high affinity peptide binding sites. These findings are qualitatively similar to those obtained by Snyderman and colleagues who studied the effect of guanine nucleotides on binding of fMet-Le~-[~H]Phe to membranes of human peripheral neutrophils [20]. However, there are important quantitative differences between the two systems.…”

“…Thus, increasing the amount of MgCI2 added to the binding reaction from 0 to 1.5 mM and 5 mM MgC12 resulted in a 4.4-fold and 8.9-fold increase in the number of high affinity receptors, respectively, whereas the number of low affinity receptors appeared to remain largely unaltered. Binding of formyl peptides to membranes prepared from human peripheral neutrophils has previously been noted to be modified by guanine nucleotides [20]. We therefore determined the role of guanine nucleotides in regulating the Mg2+-stimulated binding of fMet-Le~-[~H]Phe to HL 60 membranes.…”

Role of G protein-coupled receptors in inflammation

“…A number of small molecules was discovered in the 60's and 70's, including activated complement C5a and N-formylated peptides of bacterial origin [2][3][4] . Evidence that these "classic chemoattractants" act on GPCRs first came from the observation that pertussis toxin (PTX) could alter the binding affinity of chemotactic formyl peptides [5] , a characteristic feature of certain GPCR ligands [6] . Working independently, two laboratories reported that PTX could block formyl peptide (eg, fMet-Leu-Phe, fMLF)-induced neutrophil functions through ADP-ribosylation of the Gi class of heterotrimeric G proteins [7,8] .…”

The superoxide‐generating oxidase of phagocytic cells

“…Binding studies with neutrophils (Koo et al, 1982) and macrophages (Snyderman et al, 1984) demonstrated a single class of binding sites, with a Kd close to 20nM. Upon cell disruption, high-affinity receptor sites with a Kd of 1-2 nM were unmasked (Koo et al, 1982;Mackin et al, 1982;Snyderman et al, 1984; for review, see Snyderman et al, 1986 and.…”