Expression of Fap amyloids in Pseudomonas aeruginosa,P. fluorescens, and P. putida results in aggregation and increased biofilm formation

Abstract

The fap operon, encoding functional amyloids in Pseudomonas (Fap), is present in most pseudomonads, but so far the expression and importance for biofilm formation has only been investigated for P. fluorescens strain UK4. In this study, we demonstrate the capacity of P. aeruginosa PAO1, P. fluorescens Pf-5, and P. putida F1 to express Fap fibrils, and investigated the effect of Fap expression on aggregation and biofilm formation. The fap operon in all three Pseudomonas species conferred the ability to express Fap fibrils as shown using a recombinant approach. This Fap overexpression consistently resulted in highly aggregative phenotypes and in increased biofilm formation. Detailed biophysical investigations of purified fibrils confirmed FapC as the main fibril monomer and supported the role of FapB as a minor, nucleating constituent as also indicated by bioinformatic analysis. Bioinformatics analysis suggested FapF and FapD as a potential β-barrel membrane pore and protease, respectively. Manipulation of the fap operon showed that FapA affects monomer composition of the final amyloid fibril, and that FapB is an amyloid protein, probably a nucleator for FapC polymerization. Our study highlights the fap operon as a molecular machine for functional amyloid formation.

Knockout of Extracytoplasmic Function Sigma Factor ECF-10 Affects Stress Resistance and Biofilm Formation in Pseudomonas putida KT2440

“…Biofilms represent a common microbial life strategy, allowing bacteria to stay in a protected environment and survive even harsh conditions (47). The biofilm matrix displays a barrier that confers tolerance against various antimicrobial agents as well as the host immune response (48,49).…”

Amyloid Structures as Biofilm Matrix Scaffolds

“…Examples of extracellular fibers with structural and functional similarity to curli fimbriae of E. coli have been described in various species of Pseudomonas, including P. aeruginosa, P. fluorescens, and P. putida (36). In this case, the protein components for amyloid assembly are encoded in the fapABCDEF operon (37).…”

Biofilm Development

“…However, it is also conceivable that a specific bacterial species may form biofilm through different pathways under different conditions. P. aeruginosa, for example, can produce a number of biofilm matrix products, including the Psl, Pel, and alginate polysaccharides; extracellular DNA; and the type IV pili, Cup, CdrA, LecA, LecB, and Fap protein components (2,31,45,(51)(52)(53)(54)(55)(56)(57). It may be expected that some P. aeruginosa strains selected for under specific conditions will produce some of these biofilm matrix components in higher quantities than other strains and that the biofilm pathways will vary accordingly.…”