Characterization of a Salmonella Transcription Factor-DNA Complex and Identification of the Inducer by Native Mass Spectrometry.

Autor: Szkoda BE; Department of Chemistry and Biochemistry, The Ohio State University, Columbus, OH 43210, USA; The Ohio State Biochemistry Program, The Ohio State University, Columbus, OH 43210, USA., Di Capua A; Department of Chemistry and Biochemistry, The Ohio State University, Columbus, OH 43210, USA; Resource for Native Mass Spectrometry-Guided Structural Biology, The Ohio State University, Columbus, OH 43210, USA., Shaffer J; Department of Chemistry and Biochemistry, The Ohio State University, Columbus, OH 43210, USA., Behrman EJ; Department of Chemistry and Biochemistry, The Ohio State University, Columbus, OH 43210, USA., Wysocki VH; Department of Chemistry and Biochemistry, The Ohio State University, Columbus, OH 43210, USA; The Ohio State Biochemistry Program, The Ohio State University, Columbus, OH 43210, USA; Resource for Native Mass Spectrometry-Guided Structural Biology, The Ohio State University, Columbus, OH 43210, USA; Center for RNA Biology, The Ohio State University, Columbus, OH 43210, USA. Electronic address: wysocki.11@osu.edu., Gopalan V; Department of Chemistry and Biochemistry, The Ohio State University, Columbus, OH 43210, USA; The Ohio State Biochemistry Program, The Ohio State University, Columbus, OH 43210, USA; Center for RNA Biology, The Ohio State University, Columbus, OH 43210, USA. Electronic address: gopalan.5@osu.edu.
Jazyk: angličtina
Zdroj: Journal of molecular biology [J Mol Biol] 2022 Apr 15; Vol. 434 (7), pp. 167480. Date of Electronic Publication: 2022 Feb 14.
DOI: 10.1016/j.jmb.2022.167480
Abstrakt: FraR, a transcriptional repressor, was postulated to regulate the metabolism of the Amadori compound fructose-asparagine (F-Asn) in the foodborne pathogen Salmonella enterica. Here, the DNA- and inducer-binding affinities and stoichiometries of FraR were determined and cross-validated by electrophoretic mobility-shift assays (EMSAs) and online buffer exchange coupled to native mass spectrometry (OBE-nMS). We demonstrate the utility of OBE-nMS to characterize protein and protein-DNA complexes that are not amenable to offline exchange into volatile buffers. OBE-nMS complemented EMSAs by revealing that FraR binds to the operator DNA as a dimer and by establishing 6-phosphofructose-aspartate as the inducer that weakens DNA binding by FraR. These results provide insights into how FraR regulates the expression of F-Asn-catabolizing enzymes and add to our understanding of the intricate bacterial circuitry that dictates utilization of diverse nutrients.
Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
(Copyright © 2022 Elsevier Ltd. All rights reserved.)
Databáze: MEDLINE
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