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While GASP‐1 and GASP‐2 proteins are known to regulate myogenesis by inhibiting myostatin, their structural organization suggests a putative role as multivalent protease inhibitors controlling different protease activities. In this study, we show the noncompetitive and competitive antitrypsin activities of the full‐length GASP‐1 and GASP‐2 proteins, respectively, by using a bacterial system production and in vitro enzymatic experiments. The role of the second Kunitz domain in this functional duality is described by assessing the antitrypsin activity of GASP‐1/2 chimeric proteins. Molecular dynamics simulations support the experimental data to rationalize differences in binding modes between trypsin and the GASP‐1 and GASP‐2 second Kunitz domains. A new inhibition mechanism was evidenced for the second Kunitz domain of GASP‐2, in which the conventional cationic residue of trypsin inhibitors was substituted by the strongly interacting glutamine residue. [ABSTRACT FROM AUTHOR]

Inflammation is a major component of the vicious cycle characterizing cystic fibrosis pulmonary disease. If untreated, this inflammatory process irreversibly damages the airways, leading to bronchiectasis and ultimately respiratory failure. Antiinflammatory drugs for cystic fibrosis lung disease appear to have beneficial effects on disease parameters. These agents include oral corticosteroids and ibuprofen, as well as azithromycin, which, in addition to its antimicrobial effects, also possesses antiinflammatory properties. Inhaled corticosteroids, colchicine, methotrexate, montelukast, pentoxifylline, nutritional supplements, and protease replacement have not had a significant impact on the disease. Therapy with oral corticosteroids, ibuprofen, and fish oil is limited by adverse effects. Azithromycin appears to be safe and effective, and is thus the most promising antiinflammatory therapy available for patients with cystic fibrosis. Pharmacologic therapy with antiinflammatory agents should be started early in the disease course, before extensive irreversible lung damage has occurred. [ABSTRACT FROM AUTHOR]

We have determined effect of the oxidant peroxynitrite (ONOO-) on Ca2+-dependent matrix metalloprotease-2 (MMP-2) activity and the role of the protease on Ca2+ATPase activity in bovine pulmonary vascular smooth muscle plasma membrane under ONOO--triggered conditions. The smooth muscle plasma membrane possesses a 72-kDa protease activity in a gelatin-containing zymogram. The 72-kDa protease activity has been found to be inhibited by tissue inhibitor of metalloprotease-2 (TIMP-2), indicating that the protease is the matrix metalloprotease-2 (MMP-2). Treatment of the membrane suspension with ONOO- caused stimulation of the MMP-2 activity (as evidenced by 14C-gelatin degradation) and also increased Ca2+ATPase activity. The ONOO--triggered protease activity and the Ca2+ATPase activity were found to be inhibited by the antioxidants: vitamin E, thiourea, and mannitol. Pretreatment with catalase and superoxide dismutase did not significantly alter ONOO--stimulated MMP-2 activity and Ca2+ATPase activity, indicating that peroxide and superoxide are not present in appreciable amount in ONOO-. Under both basal and ONOO- triggered conditions, the MMP-2 activity and the Ca2+ATPase activity were also inhibited by EGTA, 1:10-phenanthroline, and TIMP-2. However, the ONOO--stimulated MMP-2 activity and the Ca2+ATPase activity were found to be insensitive to phenylmethylsulfonylfluoride, Bowman-Birk inhibitor, chymostatin, leupeptin, antipain, N-ethylmaleimide, and pepstatin. These results suggest that ONOO- caused stimulation of MMP-2 activity and that the increased MMP-2 activity subsequently played a pivotal role in stimulating Ca2+ATPase activity in bovine pulmonary vascular smooth muscle plasma membrane. [ABSTRACT FROM AUTHOR]