COMPLEXES BETWEEN SERPINS AND INACTIVE PROTEINASES ARE NOT THERMODYNAMICALLY STABLE BUT ARE RECOGNIZED BY SERPIN RECEPTORS

The serpin mechanism of action may resemble the ''standard mechanism'' described for small protein inhibitors of serine proteinases. Since t hese inhibitors are able to bind active site-modified target proteinas es, we have investigated the interactions between two serpins and thei r 3,4 -dichloroisocoumarin (DCI)-inactivated target proteinases. alpha (2)-Antiplasmin and alpha(1)-proteinase inhibitor bound stoichiometric ally to DCI-inactivated chymotrypsin (EC 3.4.21.1) and DCI-inactivated human neutrophil elastase, respectively. Similar to active proteinase s, the DCI-inactivated proteinases failed to bind complexes between se rpins and synthetic reactive site loop peptides. Thus, the abilities o f active and inactive proteinases to bind the serpins probably depend on the same structural characteristics. The thermodynamic stability of the alpha(2)-antiplasmin-DCI/chymotrypsin and alpha(1)-proteinase inh ibitor-DCI/human neutrophil elastase complexes was similar to that of virgin serpins. However, in mouse plasma elimination studies the two c omplexes were removed rapidly from the circulation, suggesting that th ey have adopted the receptor recognized conformation. Consequently, cl eavage of the reactive center peptide bond and formation of an inhibit or-acyl enzyme complex is neither obligatory to serpin-proteinase comp lex formation nor essential for the conformational change responsible for receptor mediated endocytosis.