Abstract
The ability of phenylalanine to form fibrillar nanostructures was demonstrated on multiple occasions, and such an oligomerization reaction could be the cause of cytotoxicity in patients with phenylketonuria. These findings were supported by claims that L-phenylalanine (Phe) fibrils have amyloid properties and can be detected using thioflavin T fluorescence assay. However, a part of Phe aggregation studies reported the opposite data, suggesting no amyloid structures to be formed. Due to the contradicting reports, the amyloid nature of Phe aggregates remains uncertain. In this work we tested Phe aggregation under conditions where amyloid formation was previously reported. We show the emergence of Phe aggregates with visible light optical properties that overlap with the spectra of dyes used in amyloid fibril assays, which could lead to false-positive identifications.
Highlights
Amyloid fibrils are linked to several neurodegenerative disorders, such as Alzheimer's or Parkinson's disease (Chiti & Dobson, 2006), and their formation is the result of protein aggregation into highly ordered structures (Fitzpatrick et al, 2013)
The amyloidogenic nature of Phe fibrils were supported by experimental data, showing the presence of beta-sheet conformation (Smith et al, 2008), and other amyloid-specific properties, such as thioflavin T (ThT) and Congo Red binding ability and even seeding potential (Singh et al, 2014; Shaham-Niv et al, 2015; Anand et al, 2017)
As for the 300 mM Phe solution (Fig. 3B), the emission maximum was at 485 nm and the increase in ThT fluorescence was from 26 ± 10 a.u. to 85 ± 33 a.u. (>3-time increase). With both gels we observe a similar emission maxima position as well as a substantial emission intensity growth. This maxima position correlates with the ones often seen in amyloid fibril staining with ThT (Voropai et al, 2003); it is quite obvious that agarose does not form amyloid fibrils, as it is not even an amino acid, which leads to the conclusion that such emission spectra are the result of ThT immobilization and not emission spectra of aggregated Phe before (A) and after (B) the addition of ThT
Summary
Amyloid fibrils are linked to several neurodegenerative disorders, such as Alzheimer's or Parkinson's disease (Chiti & Dobson, 2006), and their formation is the result of protein aggregation into highly ordered structures (Fitzpatrick et al, 2013). In order to find the minimum possible peptide capable of fibril formation, shorter and shorter fragments were tested, all the way down to dipeptides (Azriel & Gazit, 2001; Reches & Gazit, 2003; Gazit, 2007). It was demonstrated by Adler-Abramovich et al (2012) that even unmodified monomeric L-phenylalanine (Phe) is capable of forming fibrillar aggregates, which share similarities to amyloids. In this work we generated Phe aggregates using previously described methods and analyzed their spectroscopic profiles in order to determine whether they interacted with ThT or formed π-stacking optically active structures
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
