Abstract
During the formation of an inhibitory complex with neutrophil elastase, alpha 1 antitrypsin (alpha 1 AT) undergoes a structural rearrangement and the resulting alpha 1 AT-elastase complex becomes endowed with chemoattractant activities, mediates an increase in synthesis of alpha 1 AT, and is rapidly cleared from the circulation. In previous studies we have provided evidence that these biological activities involve the recognition of a conformation-specific domain in the alpha 1 AT molecule by a cell surface receptor on human hepatoma HepG2 cells and human monocytes. The receptor has been termed the serpin-enzyme complex (SEC) receptor because it also recognizes complex of serpins antithrombin III, alpha 1 anti-chymotrypsin, and C1 inhibitor with their cognate enzymes. Because a pentapeptide domain of alpha 1 AT (amino acids 370-374, Phe-Val-Phe-Leu-Met) is sufficient for binding to the SEC receptor and the sequence of this domain is remarkably similar to those of substance P, several other tachykinins, bombesin, and the amyloid-beta peptide, we have examined the possibility that these other ligands bind to the SEC receptor. The results indicate that substance P, several other tachykinins, and bombesin compete for binding to, and cross-linking of, the SEC receptor. The SEC receptor is distinct from the substance P receptor by several criteria. There is no substance P receptor mRNA in HepG2 cells; the SEC receptor is present in much higher density on receptor-bearing cells and binds its ligands at lower affinity than the substance P receptor; the SEC receptor is much less restricted in the specificity with which it recognizes ligand; ligands for the SEC receptor including peptide 105Y (based on alpha 1 AT sequence 359-374), alpha 1 AT-protease complexes, and bombesin do not compete for binding of substance P to a stable transfected cell line expressing the substance P receptor. Finally, we show here that the amyloid-beta peptide competes for binding to the SEC receptor but does not bind to the substance P receptor, therein raising the possibility that the SEC receptor is involved in certain biological activities, including the recently described neurotrophic and neurotoxic effects ascribed to the amyloid-beta peptide.
Highlights
During the formationof an inhibitory complex with a1AT’ plays an important role in connective tissue turnover neutrophil elastase,a1antitrypsin (a1 AT) undergoes a since it is the major physiologic inhibitor of the destructive structural rearrangement and the resulat1inAgT-elas- protease neutrophil elastase [1,2]
A synthetic peptide based on the sereceptor-bearing cells and binds its ligands at lower affinity than the substancPe receptor; the SEC receptor ismuch less restricted in thsepecificity with which it recognizes ligand; ligands for the SEC receptor including peptide 105Y,a1AT-protease complexes, and bombesin do not compete for binding of substance P to a stable transquence of this domain, 1251-peptide 105Y(SIPPEVKFNKP
01cI 30 100 300 1000 3000 10000 nM Unlabelled Competitor withaffinityconstants which are lower by2-3 orders of magnitude. These data show that substance P, several other tachykinins, bombesin, and amyloid-@peptide compete with a1AT-protease complexes for binding to theSEC receptor of HepG2 cells.substanceP, the competitor which has been most extensively studied, mediates an increase in synthesisof a1AT, a biological activity which characterizes the interaction between al AT-protease complexes and the SEC receptor
Summary
Competitionfor Binding to HepG2 Cells-First we examined receptorandthesubstance P receptor. The SECreceptor- the possibility that substanPce, amyloid-8 peptide, and bombinding domain of peptid1e05Y and a1AT is homologous besin competed for binding to the SECreceptorofHepGZ to a regionoftheamyloid-8protein(aminoacids. HepGZ cells were incubated for 2 h at recentlyimplicated in neurotrophic andneurotoxic effects 4 "Cin binding buffer with IZ5I-peptide 10i5nYsubsaturating [16]. Our data show that the amyloid-8 proteinis recog- concentrations(50 nM) in the absenceor presence of competnized by the SEC receptor and that it competes for receptor binding with an affinity similarto that of peptide105Y and a1AT-protease complexes
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