THE PRONASE RESISTANCE OF CHOLESTEROL-NUCLEATING GLYCOPROTEINS IN HUMAN BILE

Background & Aims: Many putative pronucleating proteins have been isol ated from the biliary concanavalin A (con A)-binding fraction. The pro nase resistance of the overall nucleating-promoting activity was almos t never taken into consideration, The aim of this study was to identif y the! major pronase-resistant con A-binding glycoproteins, Methods: P ronase-treated and -untreated con A-binding glycoproteins were separat ed on a Superose 12 gel permeation column (Pharmacia, Uppsala, Sweden) and tested in a crystal growth assay, Proteins were identified by ami no-terminal sequencing, Results: Con A-binding pronucleating activity eluted in two peaks on the Superose column, This activity was unaltere d after pronase treatment, Activity peak I contained too little protei n to allow amino-terminal sequencing, In activity peak II, the major p ronase-resistant con A-binding glycoproteins were identified as alpha( 1)-antitrypsin and alpha(1)-antichymotrypsin. The 130-kilodalton nucle ation promoter was identified as aminopeptidase N, but the full pronas e resistance of this protein, reported earlier, was not confirmed, Imm unoabsorptive removal of alpha(1)-antitrypsin and alpha(1)-antichymotr ypsin and immunopurification showed that only alpha(1)-antichymotrypsi n had pronucleating activity, Conclusions: The pronase resistance of t he nucleating-promoting activity of the con A-binding glycoprotein fra ction was confirmed, An important part of this activity could be attri buted to alpha(1)-antichymotrypsin, It is an acute-phase protein, as a re many other pronucleating proteins, which might indicate a general m echanism of action in gallstone formation.