Human alpha 1-proteinase inhibitor binds to extracellular matrix in vitro.
alpha 1-Proteinase inhibitor (alpha 1-PI) is the major endogenous inhibitor of human leukocyte elastase (HLE). We have employed two different methods to quantitate the binding of alpha 1-PI to extracellular matrix (ECM), composed of 51% glycoproteins and proteoglycans, 37% types I and III collagen, and 12% elastin, derived from rat heart smooth muscle cells. alpha 1-PI is tightly bound to ECM via a saturable adsorption process; the bound protein fails to dissociate from the matrix after repeated washing. Binding of alpha 1-PI is unaffected by the prior removal of ECM glycoproteins with trypsin. Binding to ECM is not decreased in the presence of high salt but is decreased at low pH. A 40-fold excess of unlabeled alpha 1-PI displaces only 50% of [125I]alpha 1-PI prebound to ECM. A 30% decrease in the levels of alpha 1-PI bound to ECM is observed after DTT washes of ECM preincubated with alpha 1-PI or when alpha 1-PI is modified with iodoacetamide prior to incubation with ECM, implying that a fraction of bound alpha 1-PI is covalently linked to ECM via disulfide bond formation. Moreover, high molecular weight complexes between [125I]alpha 1-PI and ECM components can be visualized by SDS-PAGE under nonreducing conditions but disappear upon reduction. Approximately 50% of the total alpha 1-PI bound covalently or noncovalently to ECM retains the ability to inhibit HLE-mediated ECM proteolysis. alpha 1-PI-HLE complexes bound to ECM can be visualized by SDS-PAGE following the addition of HLE to ECM that was pretreated with [125I]alpha 1-PI. alpha 1-PI from normal plasma or serum also binds to ECM with retention of immunoreactivity and partial retention of inhibitory activity. However, ECM pretreated with alpha 1-PI-deficient serum retains no HLE-inhibitory activity.
Rinehart
A R
AR
Department of Biochemistry, SUNY at Stony Brook 11794-8691.
Mallya
S
S
Simon
S R
SR
D000818:Animals; D005109:Extracellular Matrix / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D019272:Leukocyte Elastase; D009131:Muscle, Smooth, Vascular / Q000378:metabolism; D010196:Pancreatic Elastase / Q000378:metabolism; D011485:Protein Binding; D051381:Rats; D000515:alpha 1-Antitrypsin / Q000378:metabolism* / Q000502:physiology
D000515:alpha 1-Antitrypsin; D010196:Pancreatic Elastase; D019272:Leukocyte Elastase