Novel assays for assessing protein-protein interactions in Alzheimer's disease

Abstract Cell based assays more closely mimic biology than direct inhibition studies Therefore drug discovery companies are increasingly utilizing cellular screening assays Current reporter systems are unable to reliably measure in cell transient protein protein interactions Furthermore many are based upon split enzyme systems which require long incubation steps are not real time and are typically low throughput In order to improve on these existing technologies we will develop a split SUMOstar system engineered to contain a tetracysteine recognition motif for a biarsenical based fluorophores There are several advantages for using split a SUMOstar fluorescent reporter system including relatively small fusions Kda to the target proteins of interest lower non specific interaction with native proteins and the ability to take real time measurements in high throughput applications Furthermore using structural based mutagenesis we can engineer the kinetics and binding affinity of the split SUMO interaction allowing us to fine tune the reporter assay to measure transient protein protein interactions Transient protein protein interactions are key to understanding cellular processes in neurological disease states Currently these interactions are poorly understood in a cellular environment We propose to build a detection system for monitoring transient protein protein interactions in cells This will allow high throughput drug screening against new targets and enable novel research of neurological disease state pathways