Am. J. Respir. Cell Mol. Biol., Vol 13, No. 6, Dec 1995, 703-711.
Accelerated binding of secretory leukoprotease inhibitor to human leukocyte elastase mediated by single-stranded sites in DNA from tracheobronchial mucus
QL Ying and SR Simon
Department of Pathology, State University of New York at Stony Brook 11794, USA.
We have found that preparations of DNA isolated from purulent sputum
possess a novel activity which accelerates and stabilizes the binding of
human leukocyte elastase to secretory leukoprotease inhibitor, a major
endogenous antielastase in the respiratory tract. DNA in sputum is derived
from the nuclear debris of disintegrated inflammatory leukocytes, and can
attain concentrations ranging from 10(2) to 10(4) micrograms/ml, depending
on the severity of pulmonary infection and inflammation. In the presence of
23 micrograms/ml DNA, a concentration lower than those found in most
purulent sputa, the rate constant for association of secretory
leukoprotease inhibitor with elastase is increased to 1.1 x 10(8) M-1s-1,
44-fold greater than that in the absence of DNA. The equilibrium
dissociation constant for the enzyme- inhibitor complex drops to 0.7 pM,
two orders of magnitude lower than that in the absence of DNA. The
accelerating effect of DNA is further increased by thermal denaturation or
by modification with exonuclease III, while it is significantly reduced by
digestion with S1 nuclease or by binding of Escherichia coli
single-stranded DNA binding protein. The results from these experiments
indicate that the structural elements in sputum DNA that are responsible
for the accelerating effect have the characteristics of single-stranded
sites. Similar kinetic effects on elastase inhibition were also observed
with human placental DNA and genomic DNAs from a variety of other species.
These findings suggest that DNA in pulmonary secretions may participate in
antielastase defense by promoting the binding of secretory leukoprotease
inhibitor to leukocyte elastase. The results may have important
implications for use of nuclease preparations in mucolytic therapy for
cystic fibrosis.
