CsgA gatekeeper residues control nucleation but not stability of functional amyloid
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Wiley Open Access
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Wiley
2024
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| author | William P. Olsen Gaston Courtade Samuel Peña‐Díaz Madhu Nagaraj Thorbjørn V. Sønderby Frans A. A. Mulder Mette G. Malle Daniel E. Otzen |
| author_facet | William P. Olsen Gaston Courtade Samuel Peña‐Díaz Madhu Nagaraj Thorbjørn V. Sønderby Frans A. A. Mulder Mette G. Malle Daniel E. Otzen William P. Olsen Gaston Courtade Samuel Peña‐Díaz Madhu Nagaraj Thorbjørn V. Sønderby Frans A. A. Mulder Mette G. Malle Daniel E. Otzen |
| collection | Wiley Open Access |
| contents | CsgA gatekeeper residues control nucleation but not stability of functional amyloid William P. Olsen Gaston Courtade Samuel Peña‐Díaz Madhu Nagaraj Thorbjørn V. Sønderby Frans A. A. Mulder Mette G. Malle Daniel E. Otzen Protein Science AbstractFunctional amyloids, beneficial to the organism producing them, are found throughout life, from bacteria to humans. While disease‐related amyloids form by uncontrolled aggregation, the fibrillation of functional amyloid is regulated by complex cellular machinery and optimized sequences, including so‐called gatekeeper residues such as Asp. However, the molecular basis for this regulation remains unclear. Here we investigate how the introduction of additional gatekeeper residues affects fibril formation and stability in the functional amyloid CsgA from E. coli. Step‐wise introduction of additional Asp gatekeepers gradually eliminated fibrillation unless preformed fibrils were added, illustrating that gatekeepers mainly affect nucleus formation. Once formed, the mutant CsgA fibrils were just as stable as wild‐type CsgA. HSQC NMR spectra confirmed that CsgA is intrinsically disordered, and that the introduction of gatekeeper residues does not alter this ensemble. NMR‐based Dark‐state Exchange Saturation Transfer (DEST) experiments on the different CsgA variants, however, show a decrease in transient interactions between monomeric states and the fibrils, highlighting a critical role for these interactions in the fibrillation process. We conclude that gatekeeper residues affect fibrillation kinetics without compromising structural integrity, making them useful and selective modulators of fibril properties. 10.1002/pro.5178 http://creativecommons.org/licenses/by/4.0/ |
| doi_str_mv | 10.1002/pro.5178 |
| format | Artículo Open Access |
| id | wiley_oa_10_1002_pro_5178 |
| institution | Wiley Open Access |
| license_str_mv | http://creativecommons.org/licenses/by/4.0/ |
| publishDate | 2024 |
| publisher | Wiley |
| record_format | wiley_oa |
| spellingShingle | CsgA gatekeeper residues control nucleation but not stability of functional amyloid William P. Olsen Gaston Courtade Samuel Peña‐Díaz Madhu Nagaraj Thorbjørn V. Sønderby Frans A. A. Mulder Mette G. Malle Daniel E. Otzen Protein Science CsgA gatekeeper residues control nucleation but not stability of functional amyloid William P. Olsen Gaston Courtade Samuel Peña‐Díaz Madhu Nagaraj Thorbjørn V. Sønderby Frans A. A. Mulder Mette G. Malle Daniel E. Otzen Protein Science AbstractFunctional amyloids, beneficial to the organism producing them, are found throughout life, from bacteria to humans. While disease‐related amyloids form by uncontrolled aggregation, the fibrillation of functional amyloid is regulated by complex cellular machinery and optimized sequences, including so‐called gatekeeper residues such as Asp. However, the molecular basis for this regulation remains unclear. Here we investigate how the introduction of additional gatekeeper residues affects fibril formation and stability in the functional amyloid CsgA from E. coli. Step‐wise introduction of additional Asp gatekeepers gradually eliminated fibrillation unless preformed fibrils were added, illustrating that gatekeepers mainly affect nucleus formation. Once formed, the mutant CsgA fibrils were just as stable as wild‐type CsgA. HSQC NMR spectra confirmed that CsgA is intrinsically disordered, and that the introduction of gatekeeper residues does not alter this ensemble. NMR‐based Dark‐state Exchange Saturation Transfer (DEST) experiments on the different CsgA variants, however, show a decrease in transient interactions between monomeric states and the fibrils, highlighting a critical role for these interactions in the fibrillation process. We conclude that gatekeeper residues affect fibrillation kinetics without compromising structural integrity, making them useful and selective modulators of fibril properties. 10.1002/pro.5178 http://creativecommons.org/licenses/by/4.0/ |
| title | CsgA gatekeeper residues control nucleation but not stability of functional amyloid |
| topic | Protein Science |
| url | https://onlinelibrary.wiley.com/doi/10.1002/pro.5178 |