GLYCOSAMINOGLYCANS REGULATE ELASTASE INHIBITION BY OXIDIZED SECRETORYLEUKOPROTEASE INHIBITOR

Secretory leukoprotease inhibitor (SLPI) is one of the major physiolog ical inhibitors protecting respiratory epithelium from attack by exces s human leukocyte elastase (HLE), a serine protease released by neutro phils upon activation in response to inflammatory stimuli. Reaction wi th N-chlorotaurine, a major long-lived oxidant generated by activated neutrophils, oxidized all four methionine residues, but no other amino acids, in SLPI, resulting in substantial diminution of its elastase i nhibitory activity. Oxidation of the P-1' residue, Met(73), accounted for most of the diminution in activity since a site-directed mutant of SLPI with leucine at the P-1' position retained much higher residual activity after reaction with N-chlorotaurine. The diminished activity of oxidized SLPI could be almost completely restored when an iduronate -containing glycosaminoglycan, such as heparin, heparan sulfate, or de rmatan sulfate, was added to the reaction medium. Addition of a sulfat ed glucuronate-containing glycosaminoglycan, chondroitin 4- or 6-sulfa te, to the medium resulted in smaller but significant restoration of t he lost activity, whereas the effects of hyaluronic acid and keratan s ulfate were negligible. Kinetic analysis revealed that glycosaminoglyc ans greatly accelerated the association of oxidized SLPI and HLE, wher eas iduronate-containing glycosaminoglycans also stabilized the enzyme -inhibitor complex formed. Based on these findings, we suggest that ox idized SLPI is a functionally active form of the inhibitor but that ex pression of its elastase inhibitory activity is regulated by sulfated uronate-containing glycosaminoglycans. Because its methionine residues have already been oxidized, this form of SLPI is resistant to the oxi dant species that selectively attacks methionine residues in proteins. These findings indicate that SLPI may play a previously unexpected ro le in elastase inhibitory function in the lungs when significant infla mmation is present.