(323f) Improving the Understanding of Early Stage Amyloid Aggregation Using Microchannel Electrophoresis

Authors: 
Redmon, X., University of Arkansas
Hestekin, C. N., University of Arkansas
Moss, M. A., University of South Carolina
Amyloid proteins are involved in a variety of diseases including Alzheimer’s Disease (amyloid beta protein) and Type 2 Diabetes (amylin protein). Several technical challenges exist for the quantitative analysis of different sizes of amyloid proteins present during the early stages of aggregation and thus new technologies are required for enhanced protein separation. Microchannel electrophoretic techniques have emerged as powerful tools for distinction between and analysis of different protein species present at specific points in the aggregation process while photo-induced crosslinking of modified peptides (PICUP) has been used to capture the early, transient oligomeric species. A protein’s characteristics, including its tendency to aggregate, are determined largely by its primary sequence, and thus variations in an amyloid’s primary structure can be correlated to changes in its aggregation process. Here we show the results of UV-CE analysis of native amyloid beta compared to several variant forms. During native amyloid beta aggregation, three species (monomer, oligomer, and fibril) were observed. Comparing variants to native amyloid beta showed that variants possessing increased flexibility (glycine substituted for leucine) exhibited more rapid change between a larger number of species, while variants possessing decreased flexibility (alanine substituted for glycine) exhibited slower change to the same number of species.