9EYI

Cryo-EM structure of SAVED-Lon protease CCaCalpL filament bound to A4p

Summary for 9EYI

• Entry DOI: 10.2210/pdb9eyi/pdb
• EMDB information: 50054
• Descriptor: SMODS-associated and fused to various effectors domain-containing protein, A4p (5'-R(*AP*AP*AP*AP)-3') (2 entities in total)
• Functional Keywords: lon protease, saved domain, crispr, hydrolase
• Biological source: Candidatus Cloacimonas acidaminovorans; More
• Total number of polymer chains: 5
• Total formula weight: 192586.21

Authors

Tamulaitiene, G.,Sasnauskas, G.,Smalakyte, D.,Tamulaitis, G. (deposition date: 2024-04-09, release date: 2024-10-02, Last modification date: 2024-12-18)

Primary citation

Smalakyte, D.,Ruksenaite, A.,Sasnauskas, G.,Tamulaitiene, G.,Tamulaitis, G.
Filament formation activates protease and ring nuclease activities of CRISPR Lon-SAVED.
Mol.Cell, 84:4239-4255.e8, 2024

PubMed Abstract: To combat phage infection, type III CRISPR-Cas systems utilize cyclic oligoadenylates (cA) signaling to activate various auxiliary effectors, including the CRISPR-associated Lon-SAVED protease CalpL, which forms a tripartite effector system together with an anti-σ factor, CalpT, and an ECF-like σ factor, CalpS. Here, we report the characterization of the Candidatus Cloacimonas acidaminovorans CalpL-CalpT-CalpS. We demonstrate that cA binding triggers CalpL filament formation and activates it to cleave CalpT within the CalpT-CalpS dimer. This cleavage exposes the CalpT C-degron, which targets it for further degradation by cellular proteases. Consequently, CalpS is released to bind to RNA polymerase, causing growth arrest in E. coli. Furthermore, the CalpL-CalpT-CalpS system is regulated by the SAVED domain of CalpL, which is a ring nuclease that cleaves cA in a sequential three-step mechanism. These findings provide key mechanistic details for the activation, proteolytic events, and regulation of the signaling cascade in the type III CRISPR-Cas immunity.

PubMed: 39362215
DOI: 10.1016/j.molcel.2024.09.002

Experimental method

ELECTRON MICROSCOPY (2.75 Å)