Journal: Nat Commun / Year: 2023 Title: Structural analysis and architectural principles of the bacterial amyloid curli. Authors: Mike Sleutel / Brajabandhu Pradhan / Alexander N Volkov / Han Remaut / Abstract: Two decades have passed since the initial proposition that amyloids are not only (toxic) byproducts of an unintended aggregation cascade, but that they can also be produced by an organism to serve a ...Two decades have passed since the initial proposition that amyloids are not only (toxic) byproducts of an unintended aggregation cascade, but that they can also be produced by an organism to serve a defined biological function. That revolutionary idea was borne out of the realization that a large fraction of the extracellular matrix that holds Gram-negative cells into a persistent biofilm is composed of protein fibers (curli; tafi) with cross-β architecture, nucleation-dependent polymerization kinetics and classic amyloid tinctorial properties. The list of proteins shown to form so-called functional amyloid fibers in vivo has greatly expanded over the years, but detailed structural insights have not followed at a similar pace in part due to the associated experimental barriers. Here we combine extensive AlphaFold2 modelling and cryo-electron transmission microscopy to propose an atomic model of curli protofibrils, and their higher modes of organization. We uncover an unexpected structural diversity of curli building blocks and fibril architectures. Our results allow for a rationalization of the extreme physico-chemical robustness of curli, as well as earlier observations of inter-species curli promiscuity, and should facilitate further engineering efforts to expand the repertoire of curli-based functional materials.
Mass: 38033.785 Da / Num. of mol.: 3 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Pontibacter korlensis (bacteria) / Gene: PKOR_12675 / Production host: Escherichia coli BL21 (bacteria) / References: UniProt: A0A0E3UX01
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Experimental details
-
Experiment
Experiment
Method: ELECTRON MICROSCOPY
EM experiment
Aggregation state: FILAMENT / 3D reconstruction method: helical reconstruction
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Sample preparation
Component
Name: curli fiber of Pontibacteri korlensis CsgA / Type: COMPLEX Details: Sample is generated by in vitro polymerisation of Pontibacter korlensis Csga subunits Entity ID: all / Source: RECOMBINANT
Molecular weight
Experimental value: NO
Source (natural)
Organism: Pontibacter korlensis (bacteria)
Source (recombinant)
Organism: Escherichia coli (E. coli) / Strain: BL21
Buffer solution
pH: 6 / Details: 15 mM MES pH 6.0
Specimen
Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Num. of particles selected: 1817890 Details: 1000 filaments were manually boxed using e2helixboxer.py of the EMAN2 package and used as a training dataset for SPHIRE-crYOLO.
3D reconstruction
Resolution: 7.6 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 64138 / Symmetry type: HELICAL
Refine LS restraints
Refine-ID
Type
Dev ideal
Number
ELECTRONMICROSCOPY
f_bond_d
0.001
7845
ELECTRONMICROSCOPY
f_angle_d
0.421
10680
ELECTRONMICROSCOPY
f_dihedral_angle_d
10.031
2736
ELECTRONMICROSCOPY
f_chiral_restr
0.046
1161
ELECTRONMICROSCOPY
f_plane_restr
0.001
1563
+
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