1,721,032 research outputs found
Kinetic and thermodynamic analysis of leech-derived tryptase inhibitor interaction with bovine tryptase and bovine trypsin
Full text linkThe interaction of leech-derived tryptase inhibitor (LDTI) with bovine liver capsule tryptase (BLCT) and bovine trypsin has been studied using both thermodynamic and kinetic approaches. Several differences were detected: (i) the equilibrium affinity of LDTI for BLCT (K-a = 8.9 x 10(5) M-1) is about 600-fold lower than that for bovine trypsin (K-a = 5.1 x 10(8) M-1); (ii) LDTI behaves as a purely non-competitive inhibitor of BLCT, while it is a purely competitive inhibitor of bovine trypsin. These functional data are compared with those previously reported for the LDTI binding to human tryptase, where tight inhibition occurs at two of the four active sites of the tetramer (K-a = 7.1 x 10(8) M-1). Amino acid sequence alignment of BLCT, human beta II-tryptase and bovine trypsin allows us to infer some possible structural basis for the observed functional differences
The Soret circular dichroism spectrum as a probe for the heme Fe(III)-Met(80) axial bond in horse cytochrome c
No full textA spectroscopic signal sensitive to the strength of the heme iron(III)-Met(80) bond in cytochrome c represents a useful tool that will provide valuable information on the heme pocket region and redox properties of the protein. At present, the 695-nm absorption band is perhaps the simplest diagnostic signal for the axial bond; this band disappears when the Fe(III)-Met(80) bond is disrupted. From the analysis of the Soret region circular dichroism spectrum of cytochrome c under conditions that gradually induce disruption of the Fe(III)-Met(80) bond, we present evidence that the 416-nm spectral dichroic band provides independent information addressing the strength of the axial bond between Fe(III) and Met(80) in cytochrome c. Further, this study demonstrates extension of the diagnostic application to very dilute protein samples based on the useful sample concentration of 5-10 mu M vs 200-300 mu M required for 695-nm absorbance measurements. (C) 1997 Elsevier Science Inc
Mast cell tryptase, a still enigmatic enzyme
No full textTryptases constitute a subfamily of trypsin-like proteinases, stored in the mast cell secretory granules of all mammalian organisms. These enzymes are released along with other mediators into the extracellular medium upon mast cell activation/degranulation. Among the trypsin-like enzymes, tryptases are unique: they are present as active enzymes in the mast cell granules, but display activity only extracellularly, and have a specificity which is much more restricted than trypsin. Tryptases are mostly tetrameric, and in only few organisms (not in humans) are they inhibited by endogenous inhibitors in vitro. The enzymatic and molecular properties of tryptases are far better characterized that any of their plausible biological functions. On the basis of its structural and functional features it could be predicted that tryptase would not degrade a large number of proteins in vivo due to low accessibility to the tetramer central pore where the active sites face inwards. Although their biological function has not yet been clarified, tryptases seem to be involved in a number of mast cell-mediated allergic and inflammatory diseases. In particular, the involvement of tryptase in asthma, an inflammatory disease of the airways often caused by allergy, has been proposed. Here we review the present knowledge on the structure-function relationship of tryptases from different organisms, with special emphasis on human enzymes, and on their role in a variety of pathophsyiological processes
Histone-tryptase interaction: H2A N-terminal tail removal and inhibitory activity
No full textThe involvement of tryptase, the trypsin-like serine proteinase of mast cell granules, in many (patho)physiological conditions is now recognized. In vitro this enzyme is known to act as a potent growth factor for fibroblasts and epithelial cells. Moreover, a role in inflammatory diseases and in dermatological disorders characterized by increased cell turnover has been suggested for this protease. In an attempt to understand the molecular basis of tryptase activity, we have investigated the interaction in vitro between bovine tryptase and histones. Here we show that tryptase cleaves histone H2A at a specific site (Arg20-Ala21), resulting in the removal of the N-terminal flexible fragment of the molecule, Furthermore, we demonstrate that the H2A major fragment (H2A*, 109 residues) generated by hydrolysis and lacking the N-terminal domain, is a noncompetitive, reversible and highly specific inhibitor (K-i = 29 nM) of tryptase enzymatic activity. H2A* is able to inhibit the hydrolysis of a small substrate as well as the cleavage of fibronectin, a high-molecular-weight substrate of tryptase. (C) 1997 Academic Press
Direct electrochemical evidence for an equilibrium intermediate in the guanidine-induced unfolding of cytochrome c
No full textThis paper reports a voltammetric and spectroscopic investigation of the guanidine-induced unfolding of cytochrome c at neutral pH and 25 degrees C. Electrochemical data provide direct evidence for the presence of an equilibrium intermediate (form I) strictly dependent on the denaturant concentration. The midpoint potential of farm I has been determined (E(1/2) = +0.010 V vs. NHE) and its structural features defined rom analysis of the circular dichroism and absorbance spectroscopy data obtained under the same experimental conditions. From the correlation of electrochemical and spectroscopic data, we propose that the features detected by the intermediate conform to the molten globule state
Localization and interaction of bovine pancreatic trypsin inhibitor and tryptase in the granules of bovine mast cells
No full textThe interaction of bovine pancreatic trypsin inhibitor and bovine tryptase, isolated from liver capsule mast cells, was investigated. They form a complex in vitro with a K(i) of 5.6 nM at pH 8.0 and are localized within the mast cell granules, as shown by immunogold staining at the electron microscope level. In addition, double immunogold electron microscopy revealed that the inhibitor and the enzyme are present in the same granules, where they occur in clusters; this may be taken as an indication of their interaction in vivo and suggests a physiological role for bovine pancreatic trypsin inhibitor in the regulation of tryptase proteolytic activity
Monoclonal antibodies against protease inhibitors: The case of the bovine pancreatic trypsin inhibitor
No full text[No abstract available
Production and characterization of monoclonal antibodies against bovine pancreatic trypsin inhibitor
No full textIdentification of L-methionine oxidation products in tripeptides, in Met-enkephalin and in the bovine basic pancreatic trypsin inhibitor: 1H and 13C NMR study
No full textLocalization and interaction of bovine pancreatic trypsin inhibitor and tryptase in the granules of bovine mast cells
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