Structural basis for serpin inhibitor activity

Abstract

The mechanism of formation and the structures of serpin-inhibitor complexes are not completely understood, despite detailed knowledge of the structures of a number of cleaved and uncleaved inhibitor, noninhibitor, and latent serpins. It has been proposed from comparison of inhibitor and noninhibitor serpins in the cleaved and uncleaved forms that insertion of strand s4A into preexisting beta-sheet A is a requirement for serpin inhibitor activity. We have investigated the role of this strand in formation of serpin-proteinase complexes and in serpin inhibitor activity through homology modeling of wild type inhibitor, mutant substrate, and latent serpins, and of putative serpin-proteinase complexes. These models explain the high stability of the complexes and provide an understanding of substrate behavior in serpins with point mutations in s4A and of latency in plasminogen activator inhibitor I.

Interaction of subtilisins with serpins

“…Some authors have concluded that this may represent the results of serpins acting as mechanism-based inactivators (Patston et al, 1991;Gettins et al, 1992;Lawrence et al, 1995). In this case, Met 358-Ser 359 cleavage would be an absolute requirement for inhibition and the inhibited proteinase would exist with its catalytic serine acylated by the a-carbonyl of Met 358 (Lawrence et al, 1995;Wright & Scarsdale, 1995). Others have concluded that cleavage may be a result of the denaturing conditions required to visualize it and that the observed acylation (SDS-stable complex formation) is an artifact and equivalent to a trapped intermediate in the substrate hydrolysis pathway (Travis & Salvesen, 1983;Longstaff &Gaffney, 1991;Mast et al, 1991;Stein & Carrell, 1995).…”

“…Thus, considerable adaptation to the proteinase's substrate cleft may occur during complex formationsetting the serpins apart from the other serine proteinase inhibitor families. Structural and kinetic data have led some workers to propose very different inhibitory mechanisms that often include a requirement for peptide bond scission in serpins, at the P,-PI' bond, not seen in the standard mechanism inhibitors (reviewed by Wright & Scarsdale, 1995).…”

Inhibition of coagulation factors by recombinant barley serpin BSZx

“…BSZx retained nearly full activity after preincubation with 8E8 in excess and, presumably, the epitope recognized by 8E8 is not located in the loop region. More likely, partial or complete loop insertion caused by complex formation or cleavage results in a structural rearrangement disrupting the epitope [19][20][21]. This suggestion is Min Fig.…”

39 Structure function relationships of plasminogen activator inhibitor type-2 (PAI-2)

“…Hence, serpin inhibition essentially involves kinetic trapping of the protease in a stable, covalent complex that normally represents a short lived intermediate stage of the protease-substrate reaction (Hermans et al 1995;Patston et al 1991;Cooperman et al 1993;Wright & Scarsdale 1995;Stratikos & Gettins 1999). Partitioning between the substrate or inhibitory pathways is determined by the kinetics of structural changes in the serpin following cleavage of the Pl-Pl' bond (Gettins etal.…”

“…As the serpin-protease complex has not yet been crystallised, the exact nature of the interaction between serpins and their target proteases has not been directly observed (refer section 1.2.5). However, several recen studies have given valuable insights into the structure of the stable complex and along with numerous kinetic studies of the serpin-protease reaction, have provided the basis for a detailed model of the serpin inhibitory mechanism (Wright & Scarsdale 1995;Gettins et al 1996;Stratikos & Gettins 1999), as detailed below (Figure 1.4). Figure 1.4 Branched p a t h w a y m e c h a n i s m of serpin action s h o w i n g possible structures of intermediates a n d products.…”