Single-Molecule Characterization of Heterogeneous Soluble Protein Oligomers

Abstract

Soluble protein oligomers are the primary toxic species in a number of human diseases including Alzheimer's Disease, Parkinson's Disease, Type II Diabetes, cancer, and even certain viral infections. A complicating factor in establishing structure/dynamics/toxicity relationships for soluble oligomers is divergent experimental conditions, such as oligomer preparation protocols, which heavily influence experimental outcomes. Similarly, the low abundance and the structural and dynamic heterogeneity of oligomers are challenges to traditional (ensemble-averaged) biophysical/biochemical characterization. Single-molecule methods are powerful tools for probing this heterogeneity, since oligomers can be observed one at a time. We used single-molecule Förster resonance energy transfer (smFRET) to characterize the heterogeneous structures and dynamics of soluble amyloid-beta (aβ) oligomers prepared using several disparate protocols. The improved comparison between aβ oligomers generated under disparate experimental conditions will better enable correlations to be made between biophysical observables and possible pathways for mediating disease (e.g., direct membrane damage, redox activity, etc.). These correlations are a prerequisite to identifying molecular determinants of oligomer toxicity and therefore means of interfering therapeutically with their mechanisms.