Structural determinants in the stability of the serpin/proteinase complex.
Serpins inhibit serine proteinases through formation of stable 1:1 complexes. In this study we have evaluated the effects of PAI-1 neutralizing antibodies (MA) on the stability of PAI-1/proteinase complexes, partially destabilized through prolongation of the reactive center loop. MA-8H9D4, reacting with residues Arg(300), Gln(303), and Asp(305), had no effect on the stability. In contrast, MA-33H1F7 and MA-55F4C12, reacting with alpha-helix F and the turn connecting hF with s3A, affected significantly and proteinase-dependently formed PAI-1/proteinase complexes. That is, MA-33H1F7 increased the stability of both PAI-1/t-PA and u-PA complexes (7- and 3-fold, respectively) whereas MA-55F4C12 stabilized PAI-1/t-PA complexes (3-fold) but destabilized PAI-1/u-PA complexes (2-fold). It is concluded that interference with the docking site of the cognate proteinase in the preformed serpin/proteinase complex may affect the intrinsic stability. We hypothesize that this is the consequence of a decreased or increased torsion of the RCL on the catalytic triad in the proteinase.
De Taeye
B
B
Laboratory for Pharmaceutical Biology and Phytopharmacology, Faculty of Pharmaceutical Sciences, Katholieke Universiteit Leuven, E Van Evenstraat 4, Leuven BE-3000, Belgium.
Verbeke
K
K
Compernolle
G
G
Biesemans
W
W
Gils
A
A
Declerck
P J
PJ
D000911:Antibodies, Monoclonal / Q000276:immunology; D001665:Binding Sites; D046911:Macromolecular Substances; D008958:Models, Molecular; D017395:Plasminogen Activator Inhibitor 1 / Q000737:chemistry* / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D015842:Serine Proteinase Inhibitors / Q000737:chemistry* / Q000378:metabolism; D010959:Tissue Plasminogen Activator / Q000737:chemistry* / Q000378:metabolism; D014568:Urokinase-Type Plasminogen Activator / Q000737:chemistry* / Q000378:metabolism
D000911:Antibodies, Monoclonal; D046911:Macromolecular Substances; D017395:Plasminogen Activator Inhibitor 1; D015842:Serine Proteinase Inhibitors; D010959:Tissue Plasminogen Activator; D014568:Urokinase-Type Plasminogen Activator