The elicitation of an oxidative burst in phagocytes rests on the assembly of a multicomponental complex (NADPH oxidase) consisting of a membrane-associated flavocytochrome (cytochrome b 559 ), representing the redox element responsible for the NADPH-dependent reduction of oxygen to superoxide (O 2 . ), two cytosolic components (p47 phox , p67 phox ), and the small GTPase Rac (1 or 2). We found that 4-(2-aminoethyl)-benzenesulfonyl fluoride (AEBSF), an irreversible serine protease inhibitor, prevented the elicitation of O 2 . production in intact macrophages and the amphiphile-dependent activation of NADPH oxidase in a cell-free system, consisting of solubilized membrane or purified cytochrome b 559 combined with total cytosol or a mixture of recombinant p47 phox , p67 phox , and Rac1. AEBSF acted at the activation step and did not interfere with the ensuing electron flow. It did not scavenge oxygen radicals and did not affect assay reagents. Five other serine protease inhibitors (three irreversible and two reversible) were found to lack an inhibitory effect on cell-free activation of NADPH oxidase. A structure-function study of AEBSF analogues demonstrated that the presence of a sulfonyl fluoride group was essential for inhibitory activity and that compounds containing an aminoalkylbenzene moiety were more active than amidinobenzene derivatives. Exposure of the membrane fraction or of purified cytochrome b 559 , but not of cytosol or recombinant cytosolic components, to AEBSF, in the presence of a critical concentration of the activating amphiphile lithium dodecyl sulfate, resulted in a marked impairment of their ability to support cell-free NADPH oxidase activation upon complementation with untreated cytosol or cytosolic components. Kinetic analysis of the effect of varying the concentration of each of the three cytosolic components on the inhibitory potency of AEBSF indicated that this was inversely related to the concentrations of p47 phox and, to a lesser degree, p67phox . AEBSF also prevented the amphiphile-elicited translocation of p47 phox and p67 phox to the membrane. These results are interpreted as indicating that AEBSF interferes with the binding of p47 phox and/or p67 phox to cytochrome b 559 , probably by a direct effect on cytochrome b 559 .The production of reactive oxygen radicals represents the major microbicidal mechanism of phagocytes (1). Oxygen radicals are also generated, in lesser amounts, by some nonphagocytic cells, sharing the enzymatic machinery characteristic of phagocytes (2), and, under certain conditions, by plant cells (3). Interest in reactive oxygen species has also been stimulated by accumulating evidence for their involvement in the pathogenesis of diseases, ranging from respiratory distress syndrome to ischemia-reperfusion injury in several organs (4).The primordial oxygen radical produced by phagocytes is superoxide (O 2 . ). 1 It is generated, in response to appropriate stimuli, by NADPH-derived one-electron reduction of molecular oxygen, a reaction catalyzed by a membr...

BSFAB binds as a reversible competitive inhibitor to the active sites of plasmin and uPA. Additionally, BSFAB positively improved keratinocyte differentiation gene expression (transglutaminase 1). These effects were translated into improvements in SC barrier integrity clinically in subjects with dry and sensitive skin and improved their perception of having a healthy skin condition.

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2000pharmacology, medicinal chemistry, vaccines, serums pharmacology, medicinal chemistry, vaccines, serums V 1100 -2403-Amidinophenylalanine-Based Inhibitors of Urokinase.-Among 20 compounds tested, (I) is found to be the most potent inhibitor of urokinase described so far. The X-ray structure of (I) is solved in complex with trypsin, superimposed with plasminogen activator urokinase and the results suggest an unique binding mode of this inhibitor type.