Shedding light on aberrant interactions – a review of modern tools for studying protein aggregates

Abstract

The link between protein aggregation and neurodegenerative disease is well established. However, given the heterogeneity of species formed during the aggregation process, it is difficult to delineate details of the molecular events involved in generating pathological aggregates from those producing soluble monomers. As aberrant aggregates are possible pharmacological targets for the treatment of neurodegenerative diseases, the need to observe and characterise soluble oligomers has pushed traditional biophysical techniques to their limits, leading to the development of a plethora of new tools capable of detecting soluble oligomers with high precision and specificity. In this review, we discuss a range of modern biophysical techniques that have been developed to study protein aggregation, and give an overview of how they have been used to understand, in detail, the aberrant aggregation of amyloidogenic proteins associated with the two most common neurodegenerative disorders, Alzheimer's disease and Parkinson's disease.

Probing the Occurrence of Soluble Oligomers through Amyloid Aggregation Scaling Laws

“…On the other hand, there is now substantial in vivo evidence of amyloidogenic proteins also forming small soluble oligomers that spread to neighboring cells and induce downstream processes associated with neurodegeneration [1,2]. Chemical kinetics, a classical cornerstone for drug discovery [3], is hardly applicable to the study of this new and pre-eminent target [4,5,6], in part due to the lack of straightforward methods to monitor the formation of a highly heterogeneous group of species ranging from protein dimers to complex n -mers [7,8]. In contrast, extensive research has been devoted to protein aggregation kinetics based on the characteristic tinctorial properties of amyloid fibrils [9].…”

Prion protein—Semisynthetic prion protein (PrP) variants with posttranslational modifications

“…However, it can be a suitable representative only under certain conditions, including thioflavin T (ThT) fluorescence‐based following of the aggregation process. Even though with this method, insights into the characteristics and kinetics of in vitro fibril formation have been gained, just recently, the molecular basis of PrP replication was established in detail by applying a single‐molecule fluorescence methodology to characterize individual aggregates. With total internal reflection fluorescence (TIRF) microscopy, Klenerman and colleagues studied fibril fragmentation and elongation of individual murine PrP aggregates from seeded aggregation in vitro .…”

Untangling the Conformational Polymorphism of Disordered Proteins Associated With Neurodegeneration at the Single-Molecule Level

“…In addition, SMF techniques have been successfully used to characterize the complex conformational distribution and plasticity of monomeric IDPs, and molecular interactions with biological partners or aggregation inducers (Banerjee and Deniz, 2014;Lee et al, 2015). While the application of SMF approaches to uncover oligomeric states has been largely debated (Kundel et al, 2018), in this review article, we focus on the use of SMF methods to study the conformational ensemble of monomeric neurodegenerationpromoting IDPs under functional and aggregation-prone conditions. Notably, the low protein concentrations required for these techniques (in the pM or nM range) inhibit the rapid protein self-assembly.…”