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Mode of Action (MoA)
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sequesters the CBASS signalling molecule 3′3′-cGAMP.
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Evidence
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Acb4 was identified as an anti-CBASS sponge protein through a biochemical screen using phage-infected lysates tested for binding to the CBASS signal 3′3′-cGAMP. EMSA assays revealed that SPO1-infected lysates shifted radiolabeled 3′3′-cGAMP, indicating strong signal sequestration (ECO_0001807). Activity-guided fractionation and mass spectrometry (ECO_0001096) pinpointed SPO1 gp1.3 (Acb4) as the binding protein. In vitro, Acb4 suppressed Cap5 nuclease activation by sequestering 3′3′-cGAMP, blocking downstream DNA degradation. In vivo, engineered phage T4 Δacb1/Δacb2::acb4 replicated efficiently in E. coli expressing Yersinia CBASS (signaling via 3′3′-cUA), rescuing ~10⁴-fold loss in fitness seen in Acb1/Acb2-deficient T4. EMSA with infected lysates confirmed functional expression of Acb4 during infection. Acb4 failed to counteract Citrobacter CBASS (3′2′-cGAMP signaling), suggesting specificity. A 2.1 Å co-crystal structure showed Acb4 forms a tetramer that selectively binds CBASS signals with high affinity (~157 nM K_D) via nucleobase-specific binding pockets. Mutagenesis of key contact residues impaired binding, confirming their role in CBASS inhibition.
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MoA Category
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degrades or sequesters molecules utilised by host defence systems
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Subtype(s) of the defence system(s) inhibited by the protein
Defence Subtype
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Yersinia aleksiciae type I CBASS and Citrobacter portucalensis/Escherichia coli type II CBASS
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Relevant publication(s)
DOI
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10.1101/2024.12.30.630793
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Other components of the anti-defence system
Multicomponent System
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-
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Known structure in PDB
PDB ID
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9E4W
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Genome(s) encoding the protein
Protein Source
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Bacillus phage SPO1
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Defence system(s) inhibited by the protein
Defences
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CBASS
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