7yk3 | pdb_00007yk3
Crystal structure of DarTG toxin-antitoxin complex from Mycobacterium tuberculosis
Structural highlights
FunctionDART_MYCTU Toxic component of a hybrid type II/IV toxin-antitoxin (TA) system. Its toxic effect is neutralized by cognate antitoxin DarG (PubMed:27939941). ADP-ribosylates ssDNA, preferentially in the motif TTTW. Uncontrolled expression of DarT leads to ADP-ribosylation of the origin of chromosomal replication DNA in cells (in vitro the most heavily modified motifs are TTTT/A in the OriC lower strand) and growth arrest (PubMed:34408320). Is very toxic to E.coli, it cannot be expressed in E.coli (PubMed:27939941). Experiments in situ in which antitoxin DarG levels are depleted (similar to overexpression of DarT) lead to cell death; expression of wild-type DarG protein from M.tuberculosis or T.aquaticus restores growth. Cells with decreased levels of DarG are more sensitive to bedaquilline (targets respiration), DNA-damaging drugs (mitomycin C, netropsin) and transcription-targeted drugs (rifabutin and rifampicin). When DarG is depleted, a DNA-damage response is induced and mutability is increased, suggesting ADP-ribosylation of DNA is the toxic effect (PubMed:32634279).[1] [2] [3] Publication Abstract from PubMedIn the DarTG toxin-antitoxin system, the DarT toxin ADP-ribosylates single-stranded DNA (ssDNA), which stalls DNA replication and plays a crucial role in controlling bacterial growth and bacteriophage infection. This toxic activity is reversed by the N-terminal macrodomain of the cognate antitoxin DarG. DarG also binds DarT, but the role of these interactions in DarT neutralization is unknown. Here, we report that the C-terminal domain of DarG (DarG toxin-binding domain [DarG(TBD)]) interacts with DarT to form a 1:1 stoichiometric heterodimeric complex. We determined the 2.2 A resolution crystal structure of the Mycobacterium tuberculosis DarT-DarG(TBD) complex. The comparative structural analysis reveals that DarG(TBD) interacts with DarT at the DarT/ssDNA interaction interface, thus sterically occluding substrate ssDNA binding and consequently inactivating toxin by direct protein-protein interactions. Our data support a unique two-layered DarT toxin neutralization mechanism of DarG, which is important in keeping the toxin molecules in check under normal growth conditions. Structural insights into DarT toxin neutralization by cognate DarG antitoxin: ssDNA mimicry by DarG C-terminal domain keeps the DarT toxin inhibited.,Deep A, Singh L, Kaur J, Velusamy M, Bhardwaj P, Singh R, Thakur KG Structure. 2023 Apr 28:S0969-2126(23)00131-4. doi: 10.1016/j.str.2023.04.008. PMID:37167974[4] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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