Richard J. Freer

Thermal regulation of FMLP receptors on human neutrophils.

Polymorphonuclear leukocytes (PMNs) from subjects diagnosed as having iuvenile periodontitis (JP) have been categorized on the basis of their chemotactic (CTX) response to f‐met‐leu‐phe (FMLP) when assayed concurrently with PMNs from periodon‐tally healthy subjects (HP). When PMNs from JP groups demonstrating depressed CTX were assayed for lysosomal enzyme secretion (LES) in response to FMLP, there were no significant differences with respect to rate or amount. Significant differences were observed between HP and chemotactically depressed JP cells when assessed for FMLP receptor ligand binding at 23°, but not at 4°. Receptor differences observed at 23° in HP cells included an increase in amount of total binding, number of receptors, and available dispiaceable binding sites, compared with the chemotactically depressed JP PMNs, whereas the receptor affinities were similar. These data suggest that differences in FMLP receptor density in JP PMN that are chemotactically depressed may be related to processes that modulate receptor mobility and/or expression.

Characteristics of Binding of a Potent Chemotactic Formyl Tetrapeptide, Formylmethionyl-Leucyl-Phenylalanyl- Phenylalanine, to the Receptors on Rabbit Neutrophils

Binding of a potent chemotactic formyl tetrapeptide, formylmethionyl‐leucyl‐phenyl‐alanyl‐phenylalanine (fMet‐Leu‐Phe‐Phe), to the formyl peptide receptors on the rabbit neutrophil was assessed by two approaches. A tritiated preparation of fMet‐Leu‐Phe‐Phe was used for direct binding studies, whereas indirect studies comprised an assessment of the ability of the formyl tetrapeptide to competitively inhibit the binding of 35S‐labeled formylmethionyl‐leucyl‐phenylalanine. These two approaches yielded analogous results. The formyl tetrapeptide fMet‐Leu‐Phe‐Phe showed rapid and saturable binding to the same chemotactic receptors as the less potent formyl tripeptides with which it was compared. Its equilibrium‐binding pattern, however, was different: fMet‐Leu‐Phe‐Phe showed a homogeneous binding pattern, in contrast to the heterogeneity seen with the less potent compounds. The relative potencies for high‐affinity binding of the two standard formyl tripeptides and fMet‐Leu‐Phe‐Phe correlated well with their relative potencies for stimulating the biological response of degranulation; the relative potencies for low‐affinity binding correlated less well.

[13] Synthesis of chemotactic peptides

Publisher Summary This chapter discusses the synthesis of the prototype peptide N α formyl-Met-Leu-Phe-OH (fMLP). The steps are a series of repetitive deprotections of t-Boc-amino acids or peptides using trifluoroacetic acid (TFA), subsequent neutralization with N-methyl morpholine (NMM), followed by coupling of the next t-Boc-amino acid using a rapid mixed anhydride method. Although several methods are available for the formylation of peptides, consistently excellent results have been obtained using the anhydride of isovaleryl chloride and formic acid. The same general protocol is used for the synthesis of the antagonists. The prototype analog is the pentapeptide, t-Boc-Phe-Leu-Phe-Leu-Phe-OH. The final step, however, is a catalytic hydrogenolysis to yield the t-Boc-free acid form of the desired peptide. Although the synthesis for the pentapeptide is presented, the tripeptide t-Boc-Phe-Leu-Phe-OH is also fully active and only slightly less potent. No solid-phase synthesis for the antagonists has been reported.