9WUF

Cryo-EM structure of Upx

Summary for 9WUF

• Entry DOI: 10.2210/pdb9wuf/pdb
• EMDB information: 66260
• Descriptor: Restriction endonuclease (1 entity in total)
• Functional Keywords: protein structure, antiviral protein
• Biological source: Salmonella enterica subsp. enterica serovar Bredeney
• Total number of polymer chains: 1
• Total formula weight: 143024.50

Authors

Zhang, H.,Li, X. (deposition date: 2025-09-18, release date: 2026-02-11, Last modification date: 2026-08-26)

Primary citation

Zhou, R.,Liu, Y.,Zhang, Q.,Yin, Z.,Tong, J.,Zhang, C.,Zhang, L.,Li, X.,Zhao, Y.,Zhang, S.,Liu, Z.,Chen, W.,Ji, N.,Zhang, H.,Li, Z.,Yin, H.,Zuo, S.,Wei, Y.
Structural and mechanistic insights into the dual-nuclease defense protein Upx as an anti-phage system.
Nat Commun, 17:-, 2026

PubMed Abstract: Nucleic acid degradation is a common strategy for prokaryotic anti-phage systems, as exemplified by the CRISPR-Cas system. The PD-(D/E)-XK nucleases constitute a widely distributed family in these defenses. Notably, most members exhibit a single nuclease domain, while variants containing dual nuclease domains within a single polypeptide remain underexplored, and their molecular mechanisms largely obscure. Here, we biochemically and functionally study a single-protein system containing an uncharacterized PD-(D/E)-XK defense protein (Upx). As revealed by single-particle electron cryo-microscopy (cryo-EM) structure, the C-terminal domain (CTD) harboring the conserved PD-(D/E)XK catalytic core is buttressed by the N-terminal domain (NTD) and the middle domain (MD). Functional assays demonstrate that the nucleic acid binding capability of the CTD is enhanced by the MD. The NTD also displays a noncanonical, basal exonuclease activity that is auto-inhibited by MD. IP-MS experiments identify Upx-interacting phage proteins, and substrate profiling defines its physiological preferences, collectively pointing to its potential physiological targets. Notably, the phage protein gp16 was found to relieve MD-mediated inhibition of the NTD, suggesting a virus-triggered mechanism for activating Upx's dual nuclease activity. Together, these findings establish Upx as a single-protein dual-nuclease anti-phage system, expanding our understanding of bacterial immunity and informing antiviral strategy development.

PubMed: 41803127
DOI: 10.1038/s41467-026-70435-x

Experimental method

ELECTRON MICROSCOPY (3.01 Å)