title: From crystal structure to in silico epitope discovery in the Burkholderia pseudomallei flagellar hook-associated protein FlgK.
abstract: Melioidosis, caused by the Gram-negative bacterium Burkholderia pseudomallei, is a potentially fatal infection that is endemic in Southeast Asia and Northern Australia that is poorly controlled by antibiotics. Research efforts to identify antigenic components for a melioidosis vaccine have led to the identification of several proteins, including subunits forming the flagella that mediate bacterial motility, host colonization, and virulence. This study focuses on the B. pseudomallei flagellar hook-associated protein (FlgK(Bp)), and provides the first insights into the 3D structure of FlgK proteins as targets for structure-based antigen engineering. The FlgK(Bp) crystal structure (presented here at 1.8-Å resolution) reveals a multidomain fold, comprising two small β-domains protruding from a large elongated α-helical bundle core. The evident structural similarity to flagellin, the flagellar filament subunit protein, suggests that, depending on the bacterial species, flagellar hook-associated proteins are likely to show a conserved, elongated α-helical bundle scaffold coupled to a variable number of smaller domains. Furthermore, we present immune serum recognition data confirming, in agreement with previous findings, that recovered melioidosis patients produce elevated levels of antibodies against FlgK(Bp), in comparison with seronegative and seropositive healthy subjects. Moreover, we show that FlgK(Bp) has cytotoxic effects on cultured murine macrophages, suggesting an important role in bacterial pathogenesis. Finally, computational epitope prediction methods applied to the FlgK(Bp) crystal structure, coupled with in vitro mapping, allowed us to predict three antigenic regions that locate to discrete protein domains. Taken together, our results point to FlgK(Bp) as a candidate for the design and production of epitope-containing subunits/domains as potential vaccine components.
authors: Department of Biosciences, University of Milan, Italy.
authors: Louise J
authors: LJ
authors: Gourlay
authors: Rachael J
authors: RJ
authors: Thomas
authors: Claudio
authors: C
authors: Peri
authors: Oscar
authors: O
authors: Conchillo-Solé
authors: Mario
authors: M
authors: Ferrer-Navarro
authors: Arnone
authors: A
authors: Nithichanon
authors: Jordi
authors: J
authors: Vila
authors: Xavier
authors: X
authors: Daura
authors: Ganjana
authors: G
authors: Lertmemongkolchai
authors: Richard
authors: R
authors: Titball
authors: Giorgio
authors: G
authors: Colombo
authors: Martino
authors: M
authors: Bolognesi
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000907:Antibodies, Bacterial / Q000097:blood; D000942:Antigens, Bacterial / Q000737:chemistry; D001426:Bacterial Proteins / Q000737:chemistry* / Q000276:immunology / Q000502:physiology; D016957:Burkholderia pseudomallei / Q000276:immunology*; D002460:Cell Line; D003198:Computer Simulation; D018360:Crystallography, X-Ray; D000939:Epitopes / Q000737:chemistry; D006801:Humans; D008264:Macrophages / Q000276:immunology / Q000382:microbiology; D008554:Melioidosis / Q000097:blood / Q000276:immunology / Q000382:microbiology; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data
chemical_list: D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D000939:Epitopes; C088595:FlgK protein, Bacteria
keywords: Burkholderia pseudomallei; antigen; epitope discovery; flagellar hook-associated protein; structural vaccinology