Oxidized mucus proteinase inhibitor: a fairly potent neutrophil elastase inhibitor.
N-chlorosuccinimide oxidizes one of the methionine residues of mucus proteinase inhibitor with a second-order rate constant of 1.5 M-1.s-1. Cyanogen bromide cleavage and NH2-terminal sequencing show that the modified residue is methionine-73, the P'1 component of the inhibitor's active centre. Oxidation of the inhibitor decreases its neutrophil elastase inhibitory capacity but does not fully abolish it. The kinetic parameters describing the elastase-oxidized inhibitor interaction are: association rate constant kass. = 2.6 x 10(5) M-1.s-1, dissociation rate constant kdiss. = 2.9 x 10(-3) s-1 and equilibrium dissociation constant Ki = 1.1 x 10(-8) M. Comparison with the native inhibitor indicates that oxidation decreases kass. by a factor of 18.8 and increases kdiss. by a factor of 6.4, and therefore leads to a 120-fold increase in Ki. Yet, the oxidized inhibitor may still act as a potent elastase inhibitor in the upper respiratory tract where its concentration is 500-fold higher than Ki, i.e. where the elastase inhibition is pseudo-irreversible. Experiments in vitro with fibrous human lung elastin, the most important natural substrate of elastase, support this view: 1.35 microM elastase is fully inhibited by 5-6 microM oxidized inhibitor whether the enzyme-inhibitor complex is formed in the presence or absence of elastin and whether elastase is pre-adsorbed on elastin or not.
Boudier
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Laboratoire d'Enzymologie, INSERM Unité 237, Université Louis Pasteur de Strasbourg, Illkirch, France.
Bieth
J G
JG
D000595:Amino Acid Sequence; D001665:Binding Sites; D003488:Cyanogen Bromide; D006801:Humans; D007700:Kinetics; D019272:Leukocyte Elastase; D008433:Mathematics; D008715:Methionine / Q000031:analogs & derivatives / Q000494:pharmacology; D008962:Models, Theoretical; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D010196:Pancreatic Elastase / Q000037:antagonists & inhibitors* / Q000302:isolation & purification; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D053491:Proteinase Inhibitory Proteins, Secretory; D011506:Proteins / Q000737:chemistry / Q000494:pharmacology*; D015842:Serine Proteinase Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D013183:Sputum / Q000201:enzymology; D013388:Succinimides / Q000494:pharmacology
D010446:Peptide Fragments; D053491:Proteinase Inhibitory Proteins, Secretory; D011506:Proteins; D015842:Serine Proteinase Inhibitors; D013388:Succinimides; C015752:N-chlorosuccinimide; D008715:Methionine; D010196:Pancreatic Elastase; D019272:Leukocyte Elastase; D003488:Cyanogen Bromide; C013111:methionine sulfoxide