title: Identification of the docking site between a type III secretion system ATPase and a chaperone for effector cargo.
abstract: A number of Gram-negative pathogens utilize type III secretion systems (T3SSs) to inject bacterial effector proteins into the host. An important component of T3SSs is a conserved ATPase that captures chaperone-effector complexes and energizes their dissociation to facilitate effector translocation. To date, there has been limited work characterizing the chaperone-T3SS ATPase interaction despite it being a fundamental aspect of T3SS function. In this study, we present the 2.1 Å resolution crystal structure of the Salmonella enterica SPI-2-encoded ATPase, SsaN. Our structure revealed a local and functionally important novel feature in helix 10 that we used to define the interaction domain relevant to chaperone binding. We modeled the interaction between the multicargo chaperone, SrcA, and SsaN and validated this model using mutagenesis to identify the residues on both the chaperone and ATPase that mediate the interaction. Finally, we quantified the benefit of this molecular interaction on bacterial fitness in vivo using chromosomal exchange of wild-type ssaN with mutants that retain ATPase activity but no longer capture the chaperone. Our findings provide insight into chaperone recognition by T3SS ATPases and demonstrate the importance of the chaperone-T3SS ATPase interaction for the pathogenesis of Salmonella.
authors.affiliation: From the Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario L8S 4K1, Canada and the Michael G. DeGroote Institute for Infectious Disease Research, Hamilton, Ontario L8S 4K1, Canada.
authors.forename: Sarah E
authors.initials: SE
authors.lastname: Allison
authors.affiliation: From the Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario L8S 4K1, Canada and the Michael G. DeGroote Institute for Infectious Disease Research, Hamilton, Ontario L8S 4K1, Canada.
authors.forename: Brian R
authors.initials: BR
authors.lastname: Tuinema
authors.affiliation: From the Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario L8S 4K1, Canada and the Michael G. DeGroote Institute for Infectious Disease Research, Hamilton, Ontario L8S 4K1, Canada.
authors.forename: Ellen S
authors.initials: ES
authors.lastname: Everson
authors.affiliation: From the Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario L8S 4K1, Canada and the Michael G. DeGroote Institute for Infectious Disease Research, Hamilton, Ontario L8S 4K1, Canada.
authors.forename: Seiji
authors.initials: S
authors.lastname: Sugiman-Marangos
authors.affiliation: From the Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario L8S 4K1, Canada and the Michael G. DeGroote Institute for Infectious Disease Research, Hamilton, Ontario L8S 4K1, Canada.
authors.forename: Kun
authors.initials: K
authors.lastname: Zhang
authors.affiliation: From the Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario L8S 4K1, Canada and the Michael G. DeGroote Institute for Infectious Disease Research, Hamilton, Ontario L8S 4K1, Canada.
authors.forename: Murray S
authors.initials: MS
authors.lastname: Junop
authors.affiliation: From the Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario L8S 4K1, Canada and the Michael G. DeGroote Institute for Infectious Disease Research, Hamilton, Ontario L8S 4K1, Canada coombes@mcmaster.ca.
authors.forename: Brian K
authors.initials: BK
authors.lastname: Coombes
mesh_terms: D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites; D003460:Crystallization; D005260:Female; D051379:Mice; D008810:Mice, Inbred C57BL; D018832:Molecular Chaperones / Q000378:metabolism*; D062105:Molecular Docking Simulation; D014774:Virulence
chemical_list: D018832:Molecular Chaperones; D000251:Adenosine Triphosphatases
keywords: ATPase; Chaperone; Crystal Structure; Microbiology; Type III Secretion System (T3SS)