Structure of the Proteus vulgaris HigB-(HigA)2-HigB toxin-antitoxin complex
Marc A Schureck1, Tatsuya Maehigashi, Stacey J Miles
1From the Department of Biochemistry, Emory University School of Medicine, Atlanta, Georgia 30322.
Abstract:
Bacterial toxin-antitoxin (TA) systems regulate key cellular processes to promote cell survival during periods of stress. During steady-state cell growth, antitoxins typically interact with their cognate toxins to inhibit activity presumably by preventing substrate recognition. We solved two x-ray crystal structures of the Proteus vulgaris tetrameric HigB-(HigA)2-HigB TA complex and found that, unlike most other TA systems, the antitoxin HigA makes minimal interactions with toxin HigB. HigB adopts a RelE family tertiary fold containing a highly conserved concave surface where we predict its active site is located. HigA does not cover the solvent-exposed HigB active site, suggesting that, in general, toxin inhibition is not solely mediated by active site hindrance by its antitoxin. Each HigA monomer contains a helix-turn-helix motif that binds to its own DNA operator to repress transcription during normal cellular growth. This is distinct from antitoxins belonging to other superfamilies that typically only form DNA-binding motifs upon dimerization. We further show that disruption of the HigB-(HigA)2-HigB tetramer to a HigBA heterodimer ablates operator binding. Taken together, our biochemical and structural studies elucidate the novel molecular details of the HigBA TA system.
Insights
Bacterial toxin-antitoxin systems ensure cell survival. The HigBA system uniquely functions, with antitoxin HigA repressing transcription via DNA binding, not direct toxin inhibition.
Area of Science:
- Microbiology
- Structural Biology
- Molecular Biology
Background:
- Bacterial toxin-antitoxin (TA) systems are crucial for cell survival under stress.
- Antitoxins typically inhibit toxin activity by blocking substrate recognition during normal growth.
- The Proteus vulgaris HigBA TA system's mechanism is not well understood.
Purpose of the Study:
- To elucidate the molecular mechanism of the Proteus vulgaris HigBA toxin-antitoxin system.
- To determine the structural basis for HigBA system function.
Main Methods:
- X-ray crystallography was used to solve the structure of the tetrameric HigB-(HigA)2-HigB complex.
- Biochemical assays were performed to investigate DNA binding and operator repression.
- Structural analysis focused on toxin-antitoxin interactions and active site accessibility.
Main Results:
- The HigBA antitoxin (HigA) shows minimal interaction with the toxin (HigB).
- HigB possesses a RelE-like fold with a predicted active site not occluded by HigA.
- HigA monomers contain helix-turn-helix motifs for DNA binding and transcriptional repression, distinct from other TA systems.
- Disruption of the tetramer to a heterodimer abolished operator binding.
Conclusions:
- The HigBA system employs a novel mechanism where antitoxin-mediated transcriptional repression is primary.
- Toxin inhibition is not solely achieved by active site hindrance by the antitoxin.
- Structural and biochemical data reveal unique features of the HigBA TA system.
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