Assignment of a single disulphide bridge in human alpha(2)-antiplasmin: Implications for the structural and functional properties

Human alpha(2)-antiplasmin (alpha(2)AP) is a serpin involved in the regulation of blood coagulation. Most serpins, unlike smaller serine proteinase inhibitors, do not contain disulphide bridges. alpha(2)AP is an exception from this generalization and has previously been shown to contain four Cys residues organized into two disulphide bridges [Lijnen, Holmes, van Hoef, Wiman, Rodriguez and Collen (1987) fur. J. Biochem. 166, 565-574]. However, we found that alpha(2)AP incorporates iodo[C-14]acetic acid, suggesting that the protein contains reactive thiol groups. This observation prompted a re-examination of the state of the thiol groups, which revealed (i) a disulphide bridge between Cys(43) and Cys(116), (ii) that Cys(76) is bound to a cysteinyl-glycine dipeptide, and (iii) and Cys(125) exists as either a free thiol or in a mixed disulphide with another Cys residue. The disulphide identified between Cys(43) and Cys(116) appears to be conserved in orthologous proteins since the homologous Cys residues form disulphide bonds in bovine and possibly mouse alpha(2)AP. The conservation of this disulphide bridge suggests that it is important for functional aspects of alpha(2)AP. However, the structural and functional analysis described in this study does not support this conclusion.