Mar 9, 2020

Quantitating denaturation by formic acid: Imperfect repeats are essential to the stability of the functional amyloid protein FapC

bioRxiv
Line Friis Bakmann ChristensenDaniel Erik Otzen

Abstract

Bacterial functional amyloids are evolutionarily optimized to aggregate to help them fulfil their biological functions, e.g. to provide mechanical stability to biofilm. Amyloid is formed in Pseudomonas sp. by the protein FapC which contains 3 imperfect repeats connected by long linkers. Stepwise removal of these repeats slows down aggregation and increases the propensity of amyloids to fragment during the fibrillation process, but how these mechanistic properties link to fibril stability is unclear. Here we address this question. The extreme robustness of functional amyloid makes them resistant to conventional chemical denaturants, but they dissolve in formic acid (FA) at high concentrations. To quantify this, we first measured the denaturing potency of FA using 3 small acid-resistant proteins (S6, lysozyme and ubiquitin). This revealed a linear relationship between [FA] and the free energy of unfolding with a slope of m FA, as well as a robust correlation between protein residue size and m FA. We then measured the solubilisation of fibrils formed from different FapC variants (with varying number of repeats) as a function of [FA]. The resulting m FA values revealed a decline in the number of residues driving amyloid formation w...Continue Reading

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Mentioned in this Paper

Protein Aggregation, Pathological
Microbial Biofilms
Monomeric GTP-Binding Proteins
Gene Deletion
Lysozyme
Pseudomonas sp. AT
Fibril - Cell Component
Size
Formic acid
Ubiquitinated Proteins

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