help button home button
AJRCMB
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Ying, Q.-L.
Right arrow Articles by Simon, S. R.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Ying, Q.-L.
Right arrow Articles by Simon, S. R.

Am. J. Respir. Cell Mol. Biol., Volume 24, Number 1, January, 2001 83-89

Kinetics of the Inhibition of Proteinase 3 by Elafin

Qi-Long Ying and Sanford R. Simon

Departments of Pathology and Biochemistry, State University of New York at Stony Brook, Stony Brook, New York

Excessive proteolytic activity of proteinase 3 (Pr3) has been suggested to be a factor contributing to the pathogenesis of emphysema and other inflammatory disorders. We report here on the kinetics of inhibition of Pr3 by one of its major endogenous inhibitors, the 6-kD inhibitory domain of elafin. The results are consistent with a reaction mechanism in which a single elafin molecule binds a single Pr3 molecule to form a fully reversible complex. The association and dissociation rate constants, and the inhibition constant were measured to be 4.0 × 106 M-1 s-1, 1.7 × 10-3 s-1, and 4.2 × 10-10 M, respectively. Triton X-100 and dimethyl sulfoxide, which are frequently added in assay mixtures for enzymatic analysis of Pr3 activity, significantly reduced the association rate. A fraction of the total neutrophil content of Pr3 has been reported to be bound to the surface of the plasma membrane of activated and nonactivated neutrophils. In this study, we also measured the reaction rate constants of elafin with Pr3 that had been previously allowed to associate with phospholipid bilayer vesicles. Binding to the model membranes slowed down the association rate to 3.3 × 105 M-1 s-1, but the membrane-bound Pr3 and elafin formed a more stable complex, with a dissociation rate constant of 9.1 × 10-4 s-1. Based on the kinetic parameters determined here and the estimated elafin concentrations in vivo, it may be concluded that elafin plays a limited role in the regulation of proteolytic activity of Pr3 in lung secretions.

Abbreviations: N-t-butyloxycarbonyl, Boc; dimyristoyl DL-alpha -phosphatidylcholine, DMPC; sodium dimyristoyl L-alpha -phosphatidyl-DL-glycerol, DMPG; dimethylsulfoxide, DMSO; Dulbecco's modified phosphate-buffered saline, DPBS; 5,5'-dithio-bis(2-nitrobenzoic acid), DTNB; human leukocyte elastase, HLE; varepsilon  -2-picolinoyl-L-lysinyl, Lys(pic); N-alpha -methoxysuccinyl, MeO-Suc; p-nitroanilide, pNA; L-norvaline, Nva; p-nitrophenyl ester, ONp; alpha 1-proteinase inhibitor, alpha 1-PI; proteinase 3, Pr3; L-pyroglutamyl-Pro-Val-pNA, S-2484; p-chlorothiobenzyl ester, SBzl(Cl); secretory leukoprotease inhibitor, SLPI.




This article has been cited by other articles:


Home page
 J. Leukoc. Biol.Home page
E. Pezzato, M. Dona, L. Sartor, I. Dell'Aica, R. Benelli, A. Albini, and S. Garbisa
Proteinase-3 directly activates MMP-2 and degrades gelatin and Matrigel; differential inhibition by (-)epigallocatechin-3-gallate
J. Leukoc. Biol., July 1, 2003; 74(1): 88 - 94.
[Abstract] [Full Text] [PDF]

Home page
 Am. J. Respir. Cell Mol. Bio.Home page
J. Duranton and J. G. Bieth
Inhibition of Proteinase 3 by {alpha}1-Antitrypsin In Vitro Predicts Very Fast Inhibition In Vivo
Am. J. Respir. Cell Mol. Biol., July 1, 2003; 29(1): 57 - 61.
[Abstract] [Full Text] [PDF]

Home page
 Am. J. Respir. Cell Mol. Bio.Home page
Q.-L. Ying and S. R. Simon
Elastolysis by Proteinase 3 and its Inhibition by alpha 1-Proteinase Inhibitor . A Mechanism for the Incomplete Inhibition of Ongoing Elastolysis
Am. J. Respir. Cell Mol. Biol., March 1, 2002; 26(3): 356 - 361.
[Abstract] [Full Text] [PDF]


HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Proc. Am. Thorac. Soc. Am. J. Respir. Crit. Care Med.
Copyright © 2001 American Thoracic Society.