Role of disulfide exchange in alpha 1-protease inhibitor.
The major endogenous inhibitor of neutrophil elastase in the plasma, alpha 1-protease inhibitor (alpha 1-PI), has a single cysteine residue which has been shown to form mixed disulfides with a number of thiols in vitro. Under normal physiological conditions, the plasma concentrations of reduced and oxidized thiols are such that a major fraction of alpha 1-PI in the circulation in vivo is in the form of mixed disulfides [Laurell, C.-B. (1979) in The Chemistry and Physiology of Human Plasma Proteins (Bing, D. H., Ed.) pp 329-341, Pergamon, New York]. We show here that the mixed disulfide between glutathione or cysteine and alpha 1-PI (alpha 1-PI-SSG or alpha 1-PI-SScys) has an intrinsic fluorescence which distinguishes it from the reduced form of alpha 1-PI. By employing the fluorescence difference, we have measured the ratio of alpha 1-PI-SH to mixed disulfide alpha 1-PI in redox buffers of different ratios of reduced to oxidized glutathione (GSH to GSSG) or reduced to oxidized cysteine (cys to cysSScys) and have calculated an equilibrium constant and redox potential of 0.74 +/- 0.08 and 8 +/- 2 mV, respectively, for the alpha 1-PI-SH/alpha 1-PI-SSG couple and of 0.32 +/- 0.02 and 29 +/- 2 mV, respectively, for the alpha 1-PI-SH/alpha 1-PI-SScys couple. We are unable to detect any change in Trp fluorescence in the complex of alpha 1-PI and elastase when the preformed complex is added to the same GSH/GSSG or cys/cysSScys redox buffers.(ABSTRACT TRUNCATED AT 250 WORDS)
Tyagi
S C
SC
Department of Medicine, University of Missouri, Columbia 65212.
Simon
S R
SR
D000577:Amides / Q000378:metabolism; D004220:Disulfides / Q000737:chemistry*; D004591:Electrophoresis, Polyacrylamide Gel; D019272:Leukocyte Elastase; D008956:Models, Chemical; D010196:Pancreatic Elastase / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D000515:alpha 1-Antitrypsin / Q000737:chemistry* / Q000502:physiology
D000577:Amides; D004220:Disulfides; D000515:alpha 1-Antitrypsin; D010196:Pancreatic Elastase; D019272:Leukocyte Elastase