A fresh look at protein aggregation in neurodegenerative diseases; new insights from mass photometry
Neurodegenerative diseases, such as Alzheimer's and Parkinson's, both involve an accumulation of protein aggregates ,. Monitoring and studying small oligomers, which are present in the early stages of disease, can be challenging, due to their transient and heterogeneous nature .
A recent paper introduces mass photometry to investigate early-stage tau protein aggregation, in solution. It addresses the limitations of traditional methods like thioflavin-T (ThT) fluorescence assay and size-exclusion chromatography (SEC), which are used in protein aggregation studies, but are not as sensitive to smaller aggregates .
To gain further insight into this research, we spoke to the first author of this study, Simanta Sarani Paul, PhD – a postdoctoral researcher at the University of Alberta.
Keep reading and register for a free webinar where Simanta will discuss how he used mass photometry to characterize tau protein oligomerization in solution, during the early stages of the aggregation process.
Introducing the first author
Dr. Simanta Sarani Paul is a postdoctoral researcher at the University of Alberta, working on the development of novel therapeutics against neurodegeneration, by studying the early stages of protein aggregation. His previous postdoctoral research at the Max Planck Institute of Biochemistry focused on the biology of virus-encoded chaperonins. Dr. Paul received his PhD from Calcutta University, studying protein folding and conformational dynamics.
Aim: To decipher the mechanism of early tau aggregation
The aim of this paper was to monitor and better understand the early stages of tau protein aggregation, particularly in the context of neurodegenerative diseases.
According to Simanta, targeting early-stage aggregation is “crucial”. He explains: “Initially, proteins are stable and function properly. At a certain point, they become prone to aggregation. This is the point where the trouble begins. One problematic protein induces other proteins to join in and aggregate, leading to a spreading effect. Once the protein aggregation process starts, it becomes difficult to stop it. But if we can identify the problematic protein responsible for initiating aggregation and understand how it recruits others, we have a chance to intervene.”
Mass photometry reveals oligomerization dynamics in tau aggregation
The authors propose using mass photometry as an alternative approach to SEC and ThT fluorescence assay, to quantify the different populations of tau oligomers in the early stages of protein aggregation .
Simanta says: “Once protein aggregation reaches the point detectable by ThT fluorescence assay and other methods, it becomes challenging to halt the process. Mass photometry was the method of choice for our study, since it can quickly quantify the relative abundance of all the tau oligomers and show how these populations change over time, from the beginning of the aggregation process”.
The mass photometry findings reveal key insights into tau aggregation:
In the absence of aggregation inducers, tau exists in a stable equilibrium between monomers, dimers, and trimers.
When aggregation is induced (using Congo red), mass photometry shows that tau oligomerization changes over time. Higher-order oligomers start forming, including large ones (up to MDa range). The oligomer populations evolve with distinct shifts in the mass distribution, demonstrating the dynamic nature of aggregation (Fig. 1) .
Simanta comments: “By employing mass photometry, we can observe for the first time how proteins interact to form early-stage oligomers - from monomers and dimers to higher-order structures. Our research, using tau protein, has shown how these early-stage interactions accelerate the aggregation process. After a certain period, these aggregated structures further evolve into fibers.”
Fig. 1 Tracking Tau aggregation with mass photometry. (A) When Congo red is introduced, tau protein starts to aggregate, causing a rapid increase in optical density due to light scattering after approximately 40 minutes. (B) Immediately after adding Congo red, the mass distribution looks the same as without the inducer. (C) After 35 minutes of incubation, the prevalence of tau monomers decreases and mass peaks corresponding to tau tetramers and pentamers start to emerge. (D) The populations of tau protein oligomers change over time after aggregation is induced. (E) Binding events as shown in the acquisition software of the mass photometer. Figure source: 
Looking ahead: New drug targets, new therapeutic interventions
While currently focusing on the tau protein, Simanta and his colleagues plan to expand their investigations to other neurotoxic proteins, such as huntingtin, α-synuclein, and TDP-43, as well as tau mutants. The objective is to ascertain whether "the observed mechanism of tau protein aggregation follows a similar pattern in other proteins too", drawing on the commonalities they have observed in their preliminary research.
Simanta also underlines the potential therapeutic applications of this research. The aim is to find effective inhibitors for early-stage aggregation, which, if successful, could mitigate the toxic effects linked to late-stage aggregates. For this, they need robust mathematical models, which will incorporate data from all the aggregation stages.
Simanta explains: “Currently, we are working on integrating learnings from mass photometry with those from ThT experiments. If we can develop a robust mathematical model that can accurately predict the early and late stages of aggregation, it would provide a comprehensive understanding - and that's our ultimate goal.”
Many thanks to Simanta for sharing his experience with mass photometry! To learn more about how mass photometry can be used in protein studies, check out the resources below.
Learn how mass photometry can help assess changes in antibody behavior under various stress conditions. Discover how it can be used to analyze the fragmentation and aggregation of therapeutic antibodies in response to stressors like light, pH variations, and peroxide exposure.
Here you can learn how mass photometry can be used to measure protein oligomerization in different experimental conditions – and even detect rare oligomeric species.
In this webinar, you will learn how the structure of lipoprotein lipase (LPL) – an enzyme involved in lipid metabolism – was solved and how its oligomerization behaviors were studied with cutting-edge bioanalytical methods. You will discover how mass photometry can complement cryoEM data – determining the dynamic oligomerization states of LPL in solution, at varying concentrations and in the presence of additives.
 Paul, S. S., Lyons, A., Kirchner, R. & Woodside, M. T. Quantifying Oligomer Populations in Real Time during Protein Aggregation Using Single-Molecule Mass Photometry. ACS Nano (2022) doi:10.1021/acsnano.2c05739.
 Wilson, D. M. et al. Hallmarks of neurodegenerative diseases. Cell 186, 693–714 (2023). doi: 10.1016/j.cell.2022.12.032