Protein fibrillation from another small angle: Sample preparation and SAXS data collection.

Abstract

Protein fibrillation associates with several chronic, progressive, and fatal disorders, counting well-known maladies as Parkinson's, Alzheimer's, and Huntington's disease. The fibrillation process includes structural changes and aggregation of the disease specific protein, resulting in a mixture of different structural states covering nm to μm scale in varying volume fractions. SAXS uniquely enables structural investigations of such evolving mixtures but requires that the underlying main data collection experiment is carefully prepared. In this chapter, we provide very detailed instructions on how to plan and perform such protein fibrillation experiments, both before and during the SAXS data collection. The chapter is based on our own experience mainly using high-end synchrotron radiation facilities for the data collection but can equally well be applied on state-of-the-art laboratory based SAXS instruments. We accumulate the know-how from our group, established via the study of different amyloid-like proteins, applying fibrillation either in batch or in plate reader, with or without known process quenching conditions.