Molecular basis of the Druantia anti-phage defense system

Hu, H.; Popp, P.; Rutbeek, N. R.; Klein-Sousa, V.; Lopez- Mendez, B.; Roa-Eguiara, A.; Piel, D.; Pape, T. H.; Sofos, N. H.; Hendriks, I. A.; Olsen, J. V.; Songailiene, I.; Harms, A.; Erhardt, M.; Taylor, N. M. I.

doi:10.64898/2026.05.12.722862

bioRxiv · 10.64898/2026.05.12.722862

Molecular basis of the Druantia anti-phage defense system

Hu, H.·Popp, P.·Rutbeek, N. R.·Klein-Sousa, V.·Lopez- Mendez, B.·Roa-Eguiara, A.·Piel, D.·Pape, T. H.·Sofos, N. H.·Hendriks, I. A.·Olsen, J. V.·Songailiene, I.·Harms, A.·Erhardt, M.·Taylor, N. M. I.

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Abstract

Eukaryotes and prokaryotes have evolved diverse antiviral immune systems, many containing helicase modules central to defense. Druantia are widespread bacterial anti-phage defense systems, each built around a large helicase domain-containing protein, DruE, paired with variable subunits. Here, we investigate the molecular basis of a minimal two-protein module Druantia system, DruH-E. We demonstrate that DruH-E is sufficient to confer robust anti-phage defense. DruE exists in equilibrium between a monomer and an asymmetric dimer, with dimerization required for in vivo immunity. Cryo-EM structures define DruE asymmetric dimer assembly and its dsDNA unwinding mechanism, revealing a topologically closed architecture that is specifically activated by dsDNA substrates with a 3' overhang. We further identify DruH as an ssDNA-binding protein regulated by a metabolic switch, where its activity is inhibited by ATP at physiological concentrations through direct competition with ssDNA. Supported by mass spectrometry and single-cell microscopy data, we establish key determinants of the Druantia defense system and reveal how it mediates a direct antiviral immune mechanism.

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Hu, H., Popp, P., Rutbeek, N. R., Klein-Sousa, V., Lopez- Mendez, B., Roa-Eguiara, A., Piel, D., Pape, T. H., Sofos, N. H., Hendriks, I. A., Olsen, J. V., Songailiene, I., Harms, A., Erhardt, M., Taylor, N. M. I.. 2026-05-13. Molecular basis of the Druantia anti-phage defense system. https://doi.org/10.64898/2026.05.12.722862

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