Structural Insights into Curli CsgA Cross-β Fibril Architecture Inspired Repurposing of Anti-amyloid Compounds as Anti-biofilm Agents

Abstract

These authors contributed equally to this work †Correspondence to: mlandau@technion.ac.il Curli amyloid fibrils secreted by Enterobacteriaceae mediate host cell adhesion and contribute to biofilm formation, thereby promoting bacterial resistance to environmental stressors. Here, we present crystal structures of amyloid-forming segments from the major curlin subunit, CsgA, revealing steric zipper fibrils of tightly mated β-sheets, demonstrating a structural link between curli and human pathological amyloids. We propose that these cross-β segments structure the highly robust curli amyloid core. D-enantiomeric peptides, originally developed to interfere with Alzheimer's disease-associated Amyloid-β, inhibited CsgA fibrillation and reduced biofilm formation in Salmonella typhimurium. Moreover, CsgA fibrils cross-seeded fibrillation of Amyloid-β, providing further support for the proposed structural resemblance and potential for cross-species amyloid interactions. In this study, we provide structural insights into curli formation, offer a novel strategy for disrupting amyloid-structured biofilms, and hypothesize on the formation of self-propagating prion-like species originating from a microbial source that could influence neurodegenerative diseases. SignificanceAtomic resolution structural insights into the biofilm-associated curli amyloid fibril secreted by Enterobacteriaceae revealed elements of fibrillar architecture conserved between bacterial and human amyloids. This inspired us to repurpose anti-amyloid drugs designed to target human pathological amyloids as a novel class of anti-biofilm agents. Moreover, the results provide a molecular basis for understanding interspecies cross-seeding of amyloids through the generation of prion-like agents by molecular mimicry. This raises concerns regarding human exposure to exogenous sources of amyloids, such as contaminated food and amyloid-secreting microbes. Overall, we provide a novel framework for investigating interspecies amyloid interactions at the molecular level and offer novel insights into mechanisms which may underlie the evolutionary and etiological relationships between the human and microbial amylomes.

Accurate contact-based modelling of repeat proteins predicts the structure of new repeats protein families

“…The structure of their best models is visually in agreement with our model (a direct comparison is difficult as the coordinates are not available for their models). Our model is also in agreement with the partial structure of the repeat units of CsgA published by Perov et al [38]. This model contains two parallel β-sheets with individual units situated perpendicular to the fibril axis (corresponding PDB IDs are 6G8C, 6G8D, 6G8E).…”

Molecular characterization of the N-terminal half of TasA during functional amyloid assembly and its contribution toBacillus subtilisbiofilm formation

“…ThT binding assays of purified AcTasA showed a progressive increase in the fluorescence intensity of ThT at the amyloid-specific wavelength over time in a concentrationdependent manner (Fig 2C). Moreover, this ThT binding kinetics was shown in a polymerization curve similar to that observed for WT TasA in vitro and other amyloid proteins 10,[28][29][30] . As mentioned above, the amyloid core is the part of the protein most intensely folded in the final tridimensional structure of the fiber, and therefore, it exhibits remarkable physicochemical robustness, including resistance to proteases 31 .…”

Accurate contact-based modelling of repeat proteins predicts the structure of Curlin and SPW repeats

“…8c DeBenedictis et al in 2017 presented and discussed ab initio models for the Curlin repeats family members CsgA and CsgB [38], their best models is in agreement with our model (a direct comparison is difficult as the coordinates is not available of their model). The model is furthermore confirmed by the partial structure of the repeat units of CsgA published by Perov et al [39] where they crystallize in parallel β-sheets with individual units situated perpendicular to the fibril axis (corresponding PDB IDs are 6G8C, 6G8D, 6G8E).…”