Structure determination of amyloid oligomers, the pathogenic species in Alzheimer ́s disease using fast MAS NMR and microfluidics

Project Information

Oligomers-MAS-NMR

Grant agreement ID: 101038074

DOI

10.3030/101038074

Project closed

EC signature date 31 March 2021

Start date 1 December 2021

End date 30 November 2023

Funded under

• SPREADING EXCELLENCE AND WIDENING PARTICIPATION

Total cost

€ 140 202,24

EU contribution

€ 140 202,24

Coordinated by

LATVIJAS ORGANISKAS SINTEZES INSTITUTS
Latvia

Periodic Reporting for period 1 - Oligomers-MAS-NMR (Structure determination of amyloid oligomers, the pathogenic species in Alzheimer ́s disease using fast MAS NMR and microfluidics)

Reporting period: 2021-12-01 to 2023-11-30

Summary of the context and overall objectives of the project

Neurogenerative diseases (ND) are one of the major threats to an ever-aging society. Despite this, still no efficient drugs or vaccines are available. Previously, several oligomer and fibril forming proteins, where found in the brains of people who suffered from ND. Structural characterization at the atomic level of these aggregated particles would help to push the discovery of drugs and vaccine candidates against ND. Solid-state nuclear magnetic resonance spectroscopy (ssNMR) at magic angle spinning (MAS) has evolved as a powerful tool for structural biology, especially for proteins involved in ND. However, ssNMR still has a limitation of sensitivity and resolution for site-specific studies. During this project we will address this problem by exploring: (I) cell-free synthesis schemes for novel fluorine incorporation into protein aggregates. (II) engineering of microfluidic devices to prepare oligomeric shape particles. III. Fast MAS methods to detect fluorine and proton nucleus. A combination of these methods could help broaden the boundaries of MAS ssNMR spectroscopy to aid in the development of a new type of therapeutics against ND.

Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far

We established a cost-efficient cell-free protein synthesis method in incorporate non-canonical fluorinated aliphatic amino acids. For ssNMR spectroscopy we explored in details coherent and incoherent contributions to fluorine line-widths at various MAS rates. At the moment, it was the first detailed research on this topic in the ssNMR community. We acquired a multidimensional ssNMR spectra and conducted chemical shift assignments of protein aggregates, enabling us to determine the secondary structural elements and compare it with the structures obtained by cry electron microscopy. We build a stable microfluidic device using cheap, in-house manufactured components.
The results achieved in this project were presented as oral or poster presentation in local (Riga in Latvia) and international conferences (Tallin in Estonia, Paris in France, IBCG at Boston in USA). These presentations approach a wide variety of scientist in the fields of protein biochemistry, amyloid proteins and ssNMR. This allowed us to establish new scientific collaborations in Europe.
For a general audience we presented our work in "shadowing days" for high-school students and “Researchers' night” at the hosting institute. Additionally lectures about protein biochemistry and ssNMR methods were presented for university students.

Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far)

The scientific results achieved in this project showed the new possibilities of MAS ssNMR fluorine and proton detection methods in studying proteins associated with Alzheimer’s and Parkinson’s diseases. Our advances in protein cell-free synthesis have a potential to establish new biotechnological methods for protein engineering with unnatural amino acids. Both ssNMR spectroscopy and cell-free synthesis could be applied to study other challenging and large protein aggregates, uniquely contributing to the development of new therapeutics against ND.

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