BINDING OF SPAAT, THE 44-RESIDUE C-TERMINAL PEPTIDE OF ALPHA-1-ANTITRYPSIN, TO PROTEINS OF THE EXTRACELLULAR-MATRIX

SPAAT (short piece of alpha(1)-antitrypsin [AAT]), the 44-residue C-te rminal peptide of AAT, was originally isolated from human placenta [Ni emann et al. (1992): Matrix 12:233-241]. It was shown to be a competit ive inhibitor of serine proteases [Niemann et al. (in press): Biochem Biophys Acta]. The binding of SPAAT to one or more proteins of the ext racellular matrix (ECM) was initially suggested on the basis of its re covery from tissue residues following a series of extractions designed to remove easily solubilized proteins [Niemann et al. (1992): Matrix 12:233-241]. Our binding studies with the model ECMs, Matrigel and Amg el, suggested that SPAAT might be bound by a specific collagen type as well as one or more non-collagenous ECM proteins. Individual ECM comp onents were screened for their ability to bind SPAAT. When the four co mmonly occurring fiber-farming collagens (types I, II, III, and V) wer e evaluated, type III was found to be preferred. In addition, although SPAAT bound to preformed type III collagen fibers in a concentration dependent fashion, it did not bind to type III collagen molecules unde rgoing fibril formation. This is consistent with a physiological mode of interaction between SPAAT and type III collagen in vivo. Of the non -collagenous ECM macromolecules (laminin-1, fibronectin, entactin, and heparan sulfate) tested, laminin-1 was preferred. The binding of radi olabelled SPAAT to type III collagen and laminin-1 was competitively i nhibited by unlabelled SPAAT as well as an unrelated protein, human se rum albumin (HSA), to establish binding specificity. The kinetics of t he release of the bound radiolabelled SPAAT were also examined to subs tantiate the non-covalent and reversible nature of this association. T hese results support the view that susceptible proteins of the ECM may actually be coated with SPAAT in vivo, possibly affording protection against inappropriate protease digestion. (C) 1997 Wiley-Liss, Inc.