Cryo-EM and cryo-ET reveal the molecular architecture and host interactions of mycobacteriophage Douge

J Maharana and CH Wang and LA Tsai and YT Liao and CH Yang and MC Shen and LS Macale and TN Tran and J Narsico and RJ Perez and SK Tewary and JL Wu and HY Lin and SW Chang and A Franklin and PJ Moynihan and D Jacobs-Sera and KG Freeman and GF Hatfull and TL Lowary and MC Ho, CELL REPORTS, 44, 116057 (2025).

DOI: 10.1016/j.celrep.2025.116057

Recent reports highlight the efficacy of engineered mycobacteriophages to treat non-tuberculosis mycobacterial disease. Molecular insights into mycobacteriophage architecture and host interactions could allow structure-guided phage engineering to increase efficacy and broaden host range, but such information is currently unavailable. We describe the cryoelectron microscopy (cryo-EM) structure of mycobacteriophage Douge, which contains 1,105 protein subunits assembled into a complete siphophage and is coated with glycan-binding domains for mycobacterial cell surface interactions. When filled with viral genome, the channel spanning the connector, tail, and baseplate is sealed by tape measure proteins, providing a genome gating system and requiring limited structural changes for genome ejection upon phage-host contact. Nano- meter-resolution cryoelectron tomography (cryo-ET) snapshots of phage- host interactions show that the baseplate remains attached to the mycobacterial outer membrane during viral genome ejection. This study reveals high-resolution structural details of this mycobacteriophage and its interaction with host glycans.

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