Phosphorylation as conformational switch from the native to amyloid state: Trp-cage as a protein aggregation model

József Kardos, Bence Kiss, András Micsonai, Petra Rovó, Dóra K. Menyhárd, János Kovács, Györgyi Váradi, Gábor K. Tóth, András Perczel

Research output: Contribution to journalArticle

7 Citations (Scopus)

Abstract

The 20 residue long Trp-cage miniprotein is an excellent model for both computational and experimental studies of protein folding and stability. Recently, great attention emerged to study disease-related protein misfolding, aggregation, and amyloid formation, with the aim of revealing their structural and thermodynamic background. Trp-cage is sensitive to both environmental and structure-modifying effects. It aggregates with ease upon structure destabilization, and thus it is suitable for modeling aggregation and amyloid formation. Here, we characterize the amyloid formation of several sequence modified and side-chain phosphorylated Trp-cage variants. We applied NMR, circular dichroism (CD) and Fourier transform infrared (FTIR) spectroscopies, molecular dynamics (MD) simulations, and transmission electron microscopy (TEM) in conjunction with thioflavin-T (ThT) fluorescence measurements to reveal the structural consequences of side-chain phosphorylation. We demonstrate that the native fold is destabilized upon serine phosphorylation, and the resultant highly dynamic structures form amyloid-like ordered aggregates with high intermolecular β-structure content. The only exception is the D9S(P) variant, which follows an alternative aggregation process by forming thin fibrils, presenting a CD spectrum of PPII helix, and showing low ThT binding capability. We propose a complex aggregation model for these Trp-cage miniproteins. This model assumes an additional aggregated state, a collagen triple helical form that can precede amyloid formation. The phosphorylation of a single serine residue serves as a conformational switch, triggering aggregation, otherwise mediated by kinases in cell. We show that Trp-cage miniprotein is indeed a realistic model of larger globular systems of composite folding and aggregation landscapes and helps us to understand the fundamentals of deleterious protein aggregation and amyloid formation.

Original languageEnglish
Pages (from-to)2946-2955
Number of pages10
JournalJournal of Physical Chemistry B
Volume119
Issue number7
DOIs
Publication statusPublished - Feb 19 2015

ASJC Scopus subject areas

  • Physical and Theoretical Chemistry
  • Surfaces, Coatings and Films
  • Materials Chemistry

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