Serpins flex their muscle: II. Structural insights into target peptidase recognition, polymerization, and transport functions

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 serpin-peptidase 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.

FIGURE 1.

Native serpin structures form docking (Michaelis) complexes with peptidases using exosites and other factors. A, structure of native/stressed (left) and cleaved/relaxed (right) α₁AT. β-Sheet A is in red, and the RCL is in magenta. In the cleaved or covalently complexed form of the serpin (see Fig. 2A), the RCL forms an extra strand in sheet A. Native, but not cleaved, serpins are available to form docking complexes. B, superposition of the structure of the antithrombin-thrombin-heparin ternary complex (with serpin in cyan, the RCL in orange, and thrombin in yellow) with antithrombin-fXa-pentasaccharide (with serpin in green, the RCL in magenta, and fXa in blue). The structures are superposed on the serpin. C, structure of α₂-antiplasmin (green with magenta RCL). The C-terminal region functions as an exosite for plasmin; the portion visible in electron density is in cyan, and the approximate position of a docking peptidase is shown in ghost white. D, structure of protein Z-dependent inhibitor (green with magenta RCL) in complex with protein Z (cyan). The approximate position of a docking peptidase is in ghost white.

FIGURE 2.

Serpin mechanism, folding, and misfolding. A, active serpins fold into a metastable state. Following the initial interaction with a target peptidase and RCL cleavage, the serpin undergoes a radical conformational change (RCL/s4A incorporation into β-sheet A) that culminates in peptidase inhibition via distortion of the catalytic residues. B, loop-sheet model of serpin polymerization. The RCL of one molecule is inserted into open β-sheet A of another. C, domain-swapped model of serpin polymerization. The model is based on the structure of a domain-swapped antithrombin dimer, where s5A and s4A (RCL) of a donor molecule insert into β-sheet A of a recipient. D, normal serpins may fold through a polymerogenic intermediate that is stabilized by certain mutations. The black dotted arrow indicates a gap in β-sheet A that accommodates s5A to form the s4A (RCL) exposed native form in A or the s5A and s4A domain-swapped structure in C.

FIGURE 3.

Recent serpin structures. A, structure of native thermopin (24). The serpin is in green, with the RCL in magenta (part of the RCL is not visible in electron density; the dotted line shows connectivity). The C-terminal sequence is in addition to the serpin domain and folds across the front of β-sheet A. B, structure of native tengpin (26). The serpin is in green, with the RCL in magenta and helix E/s1A in dark blue. The N-terminal sequence (orange coil) is crucial for maintaining the native metastable state. C, structure of PAI-1 in complex with the somatomedin B domain of vitronectin (orange). The serpin is in green, with the RCL in magenta and helix E/s1A in dark blue. D, structure of native TBG in complex with thyroxine (28). The serpin is in green, the RCL is in magenta, and thyroxine is in cyan spheres.