Biology subjects

Pape, T. H.

Publications and source records attributed to Pape, T. H..

3 recordsLinked to original sources

bioRxiv2026

Molecular basis of the Druantia anti-phage defense system

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.

microbiology↗

bioRxiv2025

Structure of a novel contractile injection system in subsp.

Extracellular contractile injection systems (eCISs) are phage-derived nanomachines used by bacteria to deliver effectors into target cells. Well-studied examples include the Photorhabdus asymbiotica virulence cassettes and the antifeeding prophage from Serratia entomophila, which have been engineered for heterologous cargo delivery. Recent genomic analyses identified previously uncharacterized eCIS gene clusters in the opportunistic human pathogen Salmonella enterica subspecies salamae, but their structure, function, and biotechnological potential remain unexplored. Here, we report a high-resolution cryo-electron microscopy structure of the S. enterica eCIS. Our atomic models reveal a unique sheath architecture, an expansive cage-like shell around a central spike, and an associated transmembrane hydrolase. We identify a putative effector encoded within the operon exhibiting periplasmic toxicity and provide evidence that the S. enterica eCIS deviates from canonical eCISs by interacting with the inner membrane. Guided by these structural features, we uncover a previously unannotated cluster of contractile injection systems (CISs). Together, our findings expand the known diversity of CISs structures and functions and lay the groundwork for engineering customizable protein delivery platforms.

microbiology↗

bioRxiv2025

Structural basis for selective inhibition of human GABA transporter GAT3

The astrocytic {gamma}-aminobutyric acid (GABA) transporter, GAT3, is essential for terminating GABAergic signalling in the central nervous system. Selective inhibition of GAT3 offers a potential strategy for elevating extracellular GABA levels for the treatment of neurological disorders including epilepsy. However, few potent and selective GAT3 inhibitors have been developed, and their mechanisms of inhibition remain poorly understood. Here, we present the cryo-electron microscopy structures of full-length, wild-type human GAT3 bound to a selective inhibitor, to substrate GABA, or in substrate-free state. GAT3 bound to the inhibitor or in the substrate-free state exhibits an inward-open conformation. The inhibitor binds within the intracellular permeation pathway, positioned between transmembrane helices 1, 2, 3, 6, 7, and 8. The GABA-bound GAT3 is captured in an inward-occluded state, revealing the ion coordination and substrate recognition network, including a cation-{pi} interaction between GABAs {gamma}-amino group and a phenylalanine residue in transmembrane helix 6. Our data reveal the molecular determinants for the inhibitor selectivity, and the mode of substrate binding and transport inhibition, providing blueprints for the rational design of next-generation selective GAT3 inhibitors.

molecular biology↗