title: Surface-expressed enolase contributes to the pathogenesis of clinical isolate SSU of Aeromonas hydrophila.
abstract: In this study, we demonstrated that the surface-expressed enolase from diarrheal isolate SSU of Aeromonas hydrophila bound to human plasminogen and facilitated the latter's tissue-type plasminogen activator-mediated activation to plasmin. The bacterial surface-bound plasmin was more resistant to the action of its specific physiological inhibitor, the antiprotease alpha(2)-antiplasmin. We found that immunization of mice with purified recombinant enolase significantly protected the animals against a lethal challenge dose of wild-type (WT) A. hydrophila. Minimal histological changes were noted in organs from mice immunized with enolase and then challenged with WT bacteria compared to severe pathological changes found in the infected and nonimmunized group of animals. This correlated with the smaller bacterial load of WT bacteria in the livers and spleens of enolase-immunized mice than that found in the nonimmunized controls. We also showed that the enolase gene could potentially be important for the viability of A. hydrophila SSU as we could delete the chromosomal copy of the enolase gene only when another copy of the targeted gene was supplied in trans. By site-directed mutagenesis, we altered five lysine residues located at positions 343, 394, 420, 427, and 430 of enolase in A. hydrophila SSU; the mutated forms of enolase were hyperexpressed in Escherichia coli, and the proteins were purified. Our results indicated that lysine residues at positions 420 and 427 of enolase were crucial in plasminogen-binding activity. We also identified a stretch of amino acid residues ((252)FYDAEKKEY(260)) in the A. hydrophila SSU enolase involved in plasminogen binding. To our knowledge, this is the first report of the direct involvement of surface-expressed enolase in the pathogenesis of A. hydrophila SSU infections and of any gram-negative bacteria in general.
authors.affiliation: Department of Microbiology and Immunology, Sealy Center for Vaccine Development, University of Texas Medical Branch, Galveston, Texas 77555, USA.
authors.forename: Jian
authors.initials: J
authors.lastname: Sha
authors.forename: Tatiana E
authors.initials: TE
authors.lastname: Erova
authors.forename: Rebecca A
authors.initials: RA
authors.lastname: Alyea
authors.forename: Shaofei
authors.initials: S
authors.lastname: Wang
authors.forename: Juan P
authors.initials: JP
authors.lastname: Olano
authors.forename: Vijay
authors.initials: V
authors.lastname: Pancholi
authors.forename: Ashok K
authors.initials: AK
authors.lastname: Chopra
mesh_terms: D016980:Aeromonas hydrophila / Q000201:enzymology* / Q000276:immunology / Q000302:isolation & purification / Q000472:pathogenicity*; D000818:Animals; D001426:Bacterial Proteins / Q000378:metabolism*; D001428:Bacterial Vaccines / Q000276:immunology; D001665:Binding Sites; D015169:Colony Count, Microbial; D003967:Diarrhea / Q000382:microbiology; D005341:Fibrinolysin / Q000378:metabolism; D017353:Gene Deletion; D005798:Genes, Bacterial; D020043:Genes, Essential; D016905:Gram-Negative Bacterial Infections / Q000382:microbiology; D006801:Humans; D008099:Liver / Q000382:microbiology; D051379:Mice; D050296:Microbial Viability; D020125:Mutation, Missense; D010751:Phosphopyruvate Hydratase / Q000276:immunology / Q000378:metabolism*; D010958:Plasminogen / Q000378:metabolism; D011485:Protein Binding; D013154:Spleen / Q000382:microbiology; D016019:Survival Analysis; D037521:Virulence Factors / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D001428:Bacterial Vaccines; D037521:Virulence Factors; D010958:Plasminogen; D005341:Fibrinolysin; D010751:Phosphopyruvate Hydratase