FUNCTIONS OF THE N-TERMINAL DOMAIN OF SECRETORY LEUKOPROTEASE INHIBITOR

被引:40
作者
YING, QL
KEMME, M
SIMON, SR
机构
[1] SUNY STONY BROOK, DEPT PATHOL, STONY BROOK, NY 11794 USA
[2] TH DARMSTADT, INST BIOCHEM, D-64287 DARMSTADT, GERMANY
关键词
D O I
10.1021/bi00184a013
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Secretory leukoprotease inhibitor (SLPI) comprises two homologous domains: the C-terminal domain contains the reactive site, while the function of the N-terminal domain remains unknown. In order to elucidate the function of the N-terminal domain, we studied the kinetics of reactions of human leukocyte elastase with two recombinant forms of SLPI: the full-length inhibitor and the C-terminal domain alone. The reactions of elastase with the full-length inhibitor and the C-terminal domain share the same association rate constant, 2 X 10(6) M(-1) s(-1), but the complex formed with the C-terminal domain is less stable, with a dissociation rate constant of 8 x 10(4) s(-1), 5 times higher than that of, the complex with the full-length inhibitor. The binding of the full-length inhibitor to elastase is greatly accelerated by polyanions. In the presence of submicromolar concentrations (1 mu g/mL) of heparin, the association rate constant is increased by more than 1 order of magnitude. The binding of the C-terminal domain alone to elastase shows much lower sensitivity to heparin; in the presence of 5 mu M (25 mu g/mL) heparin, association of the C-terminal domain with elastase reaches a maximum rate of 7 X 10(6) M(-1) s(-1), about 3 times higher than the rate in the absence of heparin. Similar differential effects of heparin have been observed on the reactions of alpha-chymotrypsin with the two recombinant forms of SLPI. We also found that heparin has only a small effect on the binding of elastase with elafin, an elastase-specific inhibitor homologous to the C-terminal domain of SLPI. These data reveal two previously unrecognized functions of the N-terminal domain: stabilizing the elastase-inhibitor complex and mediating the activation of the inhibitor by heparin.
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页码:5445 / 5450
页数:6
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