title: WrpA Is an Atypical Flavodoxin Family Protein under Regulatory Control of the Brucella abortus General Stress Response System.
abstract: The general stress response (GSR) system of the intracellular pathogen Brucella abortus controls the transcription of approximately 100 genes in response to a range of stress cues. The core genetic regulatory components of the GSR are required for B. abortus survival under nonoptimal growth conditions in vitro and for maintenance of chronic infection in an in vivo mouse model. The functions of the majority of the genes in the GSR transcriptional regulon remain undefined. bab1_1070 is among the most highly regulated genes in this regulon: its transcription is activated 20- to 30-fold by the GSR system under oxidative conditions in vitro. We have solved crystal structures of Bab1_1070 and demonstrate that it forms a homotetrameric complex that resembles those of WrbA-type NADH:quinone oxidoreductases, which are members of the flavodoxin protein family. However, B. abortus WrbA-related protein (WrpA) does not bind flavin cofactors with a high affinity and does not function as an NADH:quinone oxidoreductase in vitro. Soaking crystals with flavin mononucleotide (FMN) revealed a likely low-affinity binding site adjacent to the canonical WrbA flavin binding site. Deletion of wrpA (ΔwrpA) does not compromise cell survival under acute oxidative stress in vitro or attenuate infection in cell-based or mouse models. However, a ΔwrpA strain does elicit increased splenomegaly in a mouse model, suggesting that WrpA modulates B. abortus interaction with its mammalian host. Despite high structural homology with canonical WrbA proteins, we propose that B. abortus WrpA represents a functionally distinct member of the diverse flavodoxin family.
  
  
  OBJECTIVE
  Brucella abortus is an etiological agent of brucellosis, which is among the most common zoonotic diseases worldwide. The general stress response (GSR) regulatory system of B. abortus controls the transcription of approximately 100 genes and is required for maintenance of chronic infection in a murine model; the majority of GSR-regulated genes remain uncharacterized. We present in vitro and in vivo functional and structural analyses of WrpA, whose expression is strongly induced by GSR under oxidative conditions. Though WrpA is structurally related to NADH:quinone oxidoreductases, it does not bind redox cofactors in solution, nor does it exhibit oxidoreductase activity in vitro. However, WrpA does affect spleen inflammation in a murine infection model. Our data provide evidence that WrpA forms a new functional class of WrbA/flavodoxin family proteins.
authors: Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, Illinois, USA Howard Taylor Ricketts Laboratory, University of Chicago, Chicago, Illinois, USA.
authors: Julien
authors: J
authors: Herrou
authors: Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, Illinois, USA Howard Taylor Ricketts Laboratory, University of Chicago, Chicago, Illinois, USA.
authors: Daniel M
authors: DM
authors: Czyż
authors: Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, Illinois, USA Howard Taylor Ricketts Laboratory, University of Chicago, Chicago, Illinois, USA.
authors: Jonathan W
authors: JW
authors: Willett
authors: Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, Illinois, USA Howard Taylor Ricketts Laboratory, University of Chicago, Chicago, Illinois, USA.
authors: Hye-Sook
authors: HS
authors: Kim
authors: Argonne National Laboratory, Argonne, Illinois, USA.
authors: Gekleng
authors: G
authors: Chhor
authors: Argonne National Laboratory, Argonne, Illinois, USA.
authors: Gyorgy
authors: G
authors: Babnigg
authors: Argonne National Laboratory, Argonne, Illinois, USA.
authors: Youngchang
authors: Y
authors: Kim
authors: Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, Illinois, USA Howard Taylor Ricketts Laboratory, University of Chicago, Chicago, Illinois, USA Department of Microbiology, University of Chicago, Chicago, Illinois, USA scrosson@uchicago.edu.
authors: Sean
authors: S
authors: Crosson
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D002003:Brucella abortus / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D005260:Female; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology*; D006801:Humans; D008264:Macrophages / Q000382:microbiology / Q000502:physiology; D051379:Mice; D008807:Mice, Inbred BALB C; D008958:Models, Molecular; D011247:Pregnancy; D011487:Protein Conformation; D013312:Stress, Physiological / Q000502:physiology*
chemical_list: D001426:Bacterial Proteins