Functional asymmetry and chemical reactivity of CsoR family persulfide sensors.
Amino Acid Sequence
Bacterial Proteins
/ chemistry
Copper
/ chemistry
Crystallography, X-Ray
Cysteine
/ chemistry
Fluorescence Polarization
Free Radicals
/ chemistry
Gram-Positive Bacteria
/ classification
Magnetic Resonance Spectroscopy
Molecular Dynamics Simulation
Operon
Protein Conformation
Repressor Proteins
/ chemistry
Sulfides
/ chemistry
Sulfur
/ chemistry
Toluene
/ analogs & derivatives
Journal
Nucleic acids research
ISSN: 1362-4962
Titre abrégé: Nucleic Acids Res
Pays: England
ID NLM: 0411011
Informations de publication
Date de publication:
02 12 2021
02 12 2021
Historique:
accepted:
14
10
2021
revised:
12
10
2021
received:
07
09
2021
pubmed:
11
11
2021
medline:
11
1
2022
entrez:
10
11
2021
Statut:
ppublish
Résumé
CstR is a persulfide-sensing member of the functionally diverse copper-sensitive operon repressor (CsoR) superfamily. While CstR regulates the bacterial response to hydrogen sulfide (H2S) and more oxidized reactive sulfur species (RSS) in Gram-positive pathogens, other dithiol-containing CsoR proteins respond to host derived Cu(I) toxicity, sometimes in the same bacterial cytoplasm, but without regulatory crosstalk in cells. It is not clear what prevents this crosstalk, nor the extent to which RSS sensors exhibit specificity over other oxidants. Here, we report a sequence similarity network (SSN) analysis of the entire CsoR superfamily, which together with the first crystallographic structure of a CstR and comprehensive mass spectrometry-based kinetic profiling experiments, reveal new insights into the molecular basis of RSS specificity in CstRs. We find that the more N-terminal cysteine is the attacking Cys in CstR and is far more nucleophilic than in a CsoR. Moreover, our CstR crystal structure is markedly asymmetric and chemical reactivity experiments reveal the functional impact of this asymmetry. Substitution of the Asn wedge between the resolving and the attacking thiol with Ala significantly decreases asymmetry in the crystal structure and markedly impacts the distribution of species, despite adopting the same global structure as the parent repressor. Companion NMR, SAXS and molecular dynamics simulations reveal that the structural and functional asymmetry can be traced to fast internal dynamics of the tetramer. Furthermore, this asymmetry is preserved in all CstRs and with all oxidants tested, giving rise to markedly distinct distributions of crosslinked products. Our exploration of the sequence, structural, and kinetic features that determine oxidant-specificity suggest that the product distribution upon RSS exposure is determined by internal flexibility.
Identifiants
pubmed: 34755876
pii: 6424783
doi: 10.1093/nar/gkab1040
pmc: PMC8643695
doi:
Substances chimiques
Bacterial Proteins
0
Free Radicals
0
Repressor Proteins
0
Sulfides
0
persulfides
0
Toluene
3FPU23BG52
Sulfur
70FD1KFU70
Copper
789U1901C5
Cysteine
K848JZ4886
dithiol
U89B11P7SC
Types de publication
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
12556-12576Subventions
Organisme : NIGMS NIH HHS
ID : R35 GM118157
Pays : United States
Informations de copyright
© The Author(s) 2021. Published by Oxford University Press on behalf of Nucleic Acids Research.
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