Structural and Functional Characterization of Vitronectin and Ail for Host Cell Invasion by Y. pestis

Abstract

Ail (attachment invasion locus) is a membrane protein that is highly expressed on the surface of the bacterium Yersinia pestis, which is the causative agent of plague. Ail interacts with various human host factors to promote pathogenesis and bacterial survival including: (1) resistance to lysis by the human complement system, a major effector of innate immunity; (2) adhesion to human cells; (3) suppression of blood clotting; and (4) enhanced fibrinolysis. Recently, we have demonstrated that Ail also recruits Vitronectin (Vn) from human serum to the Y. pestis cell surface and promotes its degradation by the Y. pestis outer membrane protease Plasminogen activator (Pla). Vn is a 459-residue multidomain, multifunctional protein that regulates the complement, coagulation and fibrinolysis pathways. It contains binding sites for a wide range of molecules, including Heparin, Integrins and pathogenic proteins. Despite its importance, little is known about the structure of Vn, or the molecular interaction between Ail and Vn to promote Y. pestis pathogenesis. Here, we demonstrate that Vn fragments can be expressed and purified from E. coli for in vitro binding assays and NMR structural studies. The data provide molecular information about the interaction of Ail with this central human protein and help explain the functional role of Ail in Y. pestis pathogenesis.