Serpins Flex Their Muscle

Abstract

Inhibitory serpins are metastable proteins that undergo a substantial conformational rearrangement to covalently trap target peptidases. The serpin reactive center loop contributes a majority of the interactions that serpins make during the initial binding to target peptidases. However, structural studies on serpinpeptidase complexes reveal a broader set of contacts on the scaffold of inhibitory serpins that have substantial influence on guiding peptidase recognition. Structural and biophysical studies also reveal how aberrant serpin folding can lead to the formation of domain-swapped serpin multimers rather than the monomeric metastable state. Serpin domain swapping may therefore underlie the polymerization events characteristic of the serpinopathies. Finally, recent structural studies reveal how the serpin fold has been adapted for non-inhibitory functions such as hormone binding.Although amino acid sequence similarity varies from 17 to 95%, key conserved residues facilitate the folding of inhibitory serpins into a metastable conformation typically comprising three ␤-sheets, eight to nine ␣-helices, and a solvent-exposed reactive center loop (RCL) 3 (Fig. 1A) (1). The conformational "stressed-to-relaxed" transition that classical inhibitory serpins undergo upon interaction with target peptidases (Figs. 1A and 2A) has been commented on in previous reviews and will not be discussed extensively here (2-5). Suffice it to say that structural snapshots are available for native forms with the RCL fully expelled or partially (e.g. P14) 4 inserted into ␤-sheet A and for inactive forms with the RCL partially (e.g. P12) or fully (e.g. RCL-cleaved serpins, the final peptidase complex, and the intact but latent conformer) inserted into ␤-sheet A (6). Together, these data provide a comprehensive picture of the range of conformational states that the serpin scaffold adopts, as well as the structural details of the conformational rearrangement that occurs upon RCL cleavage by a target peptidase (Figs. 1A and 2A). Indeed, during peptidase inhibition, it is this latter event that triggers RCL insertion, which stabilizes the acyl-enzyme complex by transposing the enzyme and deforming its active site before deacylation occurs (4).Serpins are broadly distributed throughout all major branches of life; hence, their irreversible inhibitory mechanism must provide a distinct advantage over standard mechanism inhibitors when regulating proteolytic circuits. These concepts are discussed in the accompanying minireview (7). The main purpose of this minireview is to highlight new structural information regarding serpin-peptidase complex formation, hormone binding, and pathologic polymerization. Serpin Exosite Interactions Enhance Target Peptidase RecognitionIn 2001, the crystal structure of the Michaelis complex formed between the Manduca sexta P1 Lys serpin 1B with rat S195A trypsin was published (8). As expected, the RCL is bound in a substrate-like fashion by the peptidase, poised for attack of the P1-P1Ј bond. The core interactions inv...

Structural Basis for Recognition of Urokinase-type Plasminogen Activator by Plasminogen Activator Inhibitor-1

“…In the present Michaelis complex structure, we observed that PAI-1 RCL residues P4 -P5Ј directly contact the uPA catalytic site, accounting for about two-thirds of the total contact area. Such an interaction with a long extended PЈ region of serpin was also observed on the other Michaelis complexes paired with multispecific serpins (42), e.g. P4 -P6Ј in the antithrombin⅐S195A Factor Xa complex (14) and P4 -P5Ј in antithrombin⅐thrombin⅐heparin ternary complexes (43).…”

“…This long 37-loop permits uPA to be close to PAI-1 and to form a strong interaction that includes electrostatic and cation-bonds between the 37-loop and ␤-sheet B of PAI-1. Notably, the major involvement of PAI-1 ␤-sheet B in exosite interactions is also unique to the PAI-1⅐uPA complex: in most other serpin⅐protease Michaelis complexes, it is ␤-sheet C and the surrounding residues of serpin that mediate the interaction with the protease exosites (42). These structural features constitute the foundation of PAI-1 specificity for uPA.…”

Contribution of Conserved Lysine Residues in the α2-Antiplasmin C Terminus to Plasmin Binding and Inhibition

“…These important interactions between the ␣ 2 -antiplasmin core domain, outside the immediate vicinity of P1-P1Ј, and the active site cleft of plasmin are also likely to contribute specificity to the serpin/protease interaction. Having additional exosite interactions is not uncommon in the serpin inhibition mechanism, as it may aid in the recognition of its target protein (18,19). Together with the specific binding of the ␣ 2 -antiplasmin C terminus to plasmin kringle domains, this leads to an exquisite specificity of the interaction minimizing off-target inhibition of non-cognate proteases.…”

New Caspases’ inhibitors belonging to the serpin superfamily: A novel key control point of apoptosis in mammalian tissues

“…The sequence of the RCL defines the enzyme specificity pattern of each serpin. All inhibitory serpins are irreversible covalent "suicide" protease inhibitors forming a highly stable covalent complex with their target enzyme, a complex detectable after gel electrophoresis in denaturing conditions [9,12,13]. A large set of information together with the serpin classification are also available at the following web site: http://en.wikipedia.org/wiki/Serpins.…”