Dynamic Filament Assembly Regulates the Prolyl Aminopeptidase Activity of Plant Immune Protein DM3
- 1Korea Advanced Institute of Science and Technology
Abstract
The alpha/beta hydrolase DANGEROUS MIX 3 (DM3), a proline aminopeptidase in Arabidopsis thaliana , contributes to stress resilience through both metabolic regulation and immune signaling. Its functions are partitioned within its oligomeric structure as a trimer-of-dimers, in which the immune regulatory switch resides at the dimer interface. While the enzymatic activity of DM3 is not necessary for immune response, its activity to free prolines is critical to promote tolerance to salt and drought stress. However, how this activity is regulated has remained unclear. Here, we show that DM3 undergoes dynamic and reversible assembly into higher-order filaments in a salt-sensitive manner. The cryo-electron microscopy (EM) structure of filamentous DM3 reveals that the three-stranded helical filament arises from rearrangements of a planar hexamer into a tilted hexameric configuration. Notably, within these filaments, one of the catalytic residues is flipped out to disrupt the catalytic geometry, rendering the enzyme inactive. These findings establish filament assembly as a mechanism to sequester DM3 in an inactive state and suggest that the transition between distinct oligomeric states enables DM3 to coordinate its roles in biotic and abiotic stress responses. Significance statement Proteins with multiple interaction interfaces can assemble in different ways to regulate their function, but how this flexibility controls plant stress responses remains poorly understood. The Arabidopsis protein DM3 has dual roles as a metabolic enzyme and an immune regulator, and these functions are determined by its assembly state. Here, we show that DM3 reversibly assembles into higher-order filaments in a salt-sensitive manner. This transition disrupts its active site, switching off its enzymatic activity. Our findings reveal how structural plasticity in protein assemblies enables a single protein to coordinate multiple stress-response functions, highlighting a general mechanism for regulating protein activity in plants.
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BibTeX
@misc{kim2026dynamic,
title = {{Dynamic Filament Assembly Regulates the Prolyl Aminopeptidase Activity of Plant Immune Protein DM3}},
author = {N. Kim and W.-L. Wan and Y. Y. Tan and I.-C. Jang and E. Chae and J.-J. Song},
howpublished = {bioRxiv},
year = {2026},
month = sep,
doi = {10.64898/2026.09.03.749275},
url = {https://www.biorxiv.org/content/10.64898/2026.09.03.749275v1},
note = {Public domain},
}
RIS
TY - UNPB
TI - Dynamic Filament Assembly Regulates the Prolyl Aminopeptidase Activity of Plant Immune Protein DM3
AU - N. Kim
AU - W.-L. Wan
AU - Y. Y. Tan
AU - I.-C. Jang
AU - E. Chae
AU - J.-J. Song
PY - 2026
DA - 2026/09/03
PB - bioRxiv
DO - 10.64898/2026.09.03.749275
UR - https://www.biorxiv.org/content/10.64898/2026.09.03.749275v1
AB - The alpha/beta hydrolase DANGEROUS MIX 3 (DM3), a proline aminopeptidase in Arabidopsis thaliana , contributes to stress resilience through both metabolic regulation and immune signaling. Its functions are partitioned within its oligomeric structure as a trimer-of-dimers, in which the immune regulatory switch resides at the dimer interface. While the enzymatic activity of DM3 is not necessary for immune response, its activity to free prolines is critical to promote tolerance to salt and drought stress. However, how this activity is regulated has remained unclear. Here, we show that DM3 undergoes dynamic and reversible assembly into higher-order filaments in a salt-sensitive manner. The cryo-electron microscopy (EM) structure of filamentous DM3 reveals that the three-stranded helical filament arises from rearrangements of a planar hexamer into a tilted hexameric configuration. Notably, within these filaments, one of the catalytic residues is flipped out to disrupt the catalytic geometry, rendering the enzyme inactive. These findings establish filament assembly as a mechanism to sequester DM3 in an inactive state and suggest that the transition between distinct oligomeric states enables DM3 to coordinate its roles in biotic and abiotic stress responses. Significance statement Proteins with multiple interaction interfaces can assemble in different ways to regulate their function, but how this flexibility controls plant stress responses remains poorly understood. The Arabidopsis protein DM3 has dual roles as a metabolic enzyme and an immune regulator, and these functions are determined by its assembly state. Here, we show that DM3 reversibly assembles into higher-order filaments in a salt-sensitive manner. This transition disrupts its active site, switching off its enzymatic activity. Our findings reveal how structural plasticity in protein assemblies enables a single protein to coordinate multiple stress-response functions, highlighting a general mechanism for regulating protein activity in plants.
ER -