title: Engineering D-helix of antithrombin in alpha-1-proteinase inhibitor confers antiinflammatory properties on the chimeric serpin.
abstract: Antithrombin (AT) is a heparin-binding serpin in plasma which regulates the proteolytic activity of procoagulant proteases of the clotting cascade. In addition to being an anticoagulant, AT also exhibits antiinflammatory activities when it binds to cell surface heparan sulfate proteoglycans (HSPGs) on the endothelium via its basic residues of D-helix to elicit intracellular signalling responses. By contrast to AT, α1-proteinase inhibitor (α1-PI) is a non-heparin-binding serpin that exhibits very slow reactivity with coagulation proteases and possesses no HSPG-dependent antiinflammatory properties. To determine whether the antiinflammatory signaling specificity of AT can be transferred to α1-PI, we replaced the D-helix of human α1-PI with the corresponding sequence of human AT and expressed the chimeric serpin α1-PI/D-helix) in a bacterial expression system. High molecular weight heparin bound to α1-PI/D-helix and accelerated the inhibition of thrombin by the serpin mutant by a template mechanism reminiscent of the cofactor effect of heparin on inhibition of thrombin by AT. Like AT, α1-PI/D-helix exhibited antiinflammatory properties in both cellular and animal models. Thus, α1-PI/D-helix inhibited the barrier-disruptive effect of proinflammatory cytokines and inhibited the activation of nuclear factor-κB transcription factor in lipopolysaccharide-stimulated endothelial cells by a concentration-dependent manner. Furthermore, the chimeric serpin reduced lipopolysaccharide-mediated lethality, elicited a vascular protective effect and inhibited infiltration of activated leukocytes to the peritoneal cavity of mice in an HMGB1-mediated inflammatory model. These results suggest that grafting the D-helix of AT to α1-PI confers antiinflammatory properties on the serpin and that the chimeric serpin may have therapeutic utility for treating inflammatory disorders.
authors: L
authors: L
authors: Yang
authors: P
authors: P
authors: Dinarvand
authors: S H
authors: SH
authors: Qureshi
authors: Alireza R. Rezaie, PhD, Department of Biochemistry and Molecular Biology, St. Louis University School of Medicine, 1100 S. Grand Blvd., St. Louis, MO 63104, USA, Tel.: +1 314 977 9240, Fax: +1 314 977 9205 , E-mail: rezaiear@slu.edu.
authors: A R
authors: AR
authors: Rezaie
mesh_terms: D000818:Animals; D000991:Antithrombins / Q000502:physiology*; D001777:Blood Coagulation / Q000235:genetics; D002465:Cell Movement / Q000235:genetics; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000502:physiology*; D006493:Heparin / Q000031:analogs & derivatives / Q000378:metabolism; D006801:Humans; D007249:Inflammation / Q000276:immunology* / Q000628:therapy; D051379:Mice; D023421:Models, Animal; D009154:Mutation / Q000235:genetics; D016328:NF-kappa B / Q000378:metabolism; D015202:Protein Engineering; D017433:Protein Structure, Secondary / Q000235:genetics; D011509:Proteoglycans / Q000378:metabolism; D015398:Signal Transduction / Q000235:genetics; D000515:alpha 1-Antitrypsin / Q000235:genetics / Q000378:metabolism*
chemical_list: D000991:Antithrombins; D016328:NF-kappa B; D011509:Proteoglycans; D000515:alpha 1-Antitrypsin; C036090:heparin proteoglycan; D006493:Heparin
keywords: Antithrombin; SERPINs; coagulation inhibitors; endothelial cells; inflammation