Inhibitory activity and conformational transition of α1‐proteinase inhibitor variants

Open Access
Publisher Website
Google Scholar
Abstract
Several variants of α1-proteinase inhibitor (α1-PI) were investigated by spectroscopic methods and characterized according to their inhibitory activity. Replacement of Thr345 (P14) with Arg in α1-PI containing an Arg residue in position 358 (yielding [Thr345 Arg, Met358 Arg]α1-PI) results in complete loss of its inhibitory activity against human α-thrombin; whereas an exchange of residue Met351 (P8) by Glu ([Met351 Glu, Met358 Arg]α1-PI) does not alter activity. [Thr345 Arg, Met358 Arg]α1-PI is rapidly cleaved by thrombin, while [Met358 Arg]α1-PI and [Met351 Glu, Met358 Arg]α1-PI form stable proteinase-inhibitor complexes. The stability of [Thr345 Arg, Met358 Arg]α1-PI against guanidinium chloride denaturation is significantly enhanced compared to wild-type α1-PI, and does not change after cleavage, resembling ovalbumin, a serpin with no inhibitory activity, from which the Thr345 Arg amino acid exchange had been derived. [Met351 Glu, Met358 Arg]α1-PI and [Met358 Arg]α1-PI resemble the wild-type protein in this respect. The CD spectra of intact and cleaved α1-PI variants do not compare well with the wild-type protein, probably reflecting local structural differences. Insertion of a synthetic peptide, which corresponds to residues Thr345 Met358 of human α1-PI, leads to the formation of binary complexes with all variants having the characteristic features of the binary complex between peptide and wild-type protein.