The crystal structure of plasminogen activator inhibitor 2 at 2.0 Å resolution: implications for serpin function

Abstract

A statistical analysis of interstrand interactions indicated that the shutter region can be used to discriminate between inhibitory and non-inhibitory serpins. This analysis implied that insertion of the RCL into betasheet A up to residue P8 is important for protease inhibition and hence the structure of the complex formed between the serpin and the target protease.

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

“…In several crystal structures (inc PAI-2) the R C L is highly disordered and hence not well resolved. Taken together, these data indicate that the R C L is quite flexible and m a y alternate between a variety of conformations, the stabilisation of any particular form requiring interaction with a neighbouring molecule such as the target protease (Harrop et al 1999). …”

“…Reactive bond cleavage and subsequent strand insertion associated with the R state renders the serpin molecule permanently inactive (Wright & Searsdale 1995). Based on a statistical analysis of serpin amino acid sequences, Harrop et al (1999) suggest that the R conformation is primarily stabilised by interactions between the RCL and P-sheet A at two well conserved sites. This analysis indicated that hydrogen bonding and electrostatic interactions between P-strands s3A and s5A and the adjacent P8 and P13 residues are the principal stabilising factors in relaxed serpins.…”

“…This hydrogen bond network is further stabilised by a balance of oppositely charged residues on the RCL proximal hinge and the top of p strand s3A. (Harrop et al 1999;Elliot et al 1996).…”

“…The gap between the top of the strands is bridged by a cluster of water molecules in the structure of PAI-2. Furthermore, the sidechain of the conserved glutamate residue at P17 (Glu 364 in PAI-2) is held in position by hydrogen bonds to adjacent residues Lys 2I4 and Gln 311 (Harrop et al 1999). The importance of stabilising bonds in the proximal hinge region is further demonstrated by the impaired inhibitory activity of the oc r PI Z mutant, in which the P17 (Glu 342 ) is mutated to lysine (Jeppsson 1976).…”

“…Statistical analysis of bridging pair correlations between individual RCL residues and adjacent residues on strands 3 and 5 of p-sheet A of all known serpins (Harrop et al 1999) indicate that the S8 site (residues in P-sheet A adjacent to P8) is a reliable determinant of inhibitory activity. Experimental support for this analysis was provid by Audenaert et al (1994) who showed that mutation of the P8 residue converted PAI-1 from an inhibitor to a substrate.…”