Joydip Das
University of Houston, USA
Title: Identification of alcohol binding residues in brain proteins using electrospray ionization mass spectrometry
Biography
Biography: Joydip Das
Abstract
Alcohols bind and regulate functions of many proteins in the human brain. To develop antagonists for the alcohol addiction and alcoholism, it is necessary to identify the site of action of alcohol in these proteins. We have investigated alcohol binding sites in protein kinase C (PKC) epsilon, a serine/threonine kinase and Munc13-1, a presynaptic protein expressed predominantly in the brain. To identify alcohol binding site(s), C1 domains of PKCï¥ and Munc13-1 were photolabeled with the diazirine analogs of butanol and octanol, 3-azibutanol and 3-azioctanol, and the sites of photoincorporation were identified by MS/MS analysis using electrospray ionization mode. Azialcohols labeled Tyr-176 of PKCï¥C1A, His-248 and Tyr-250 of PKCï¥C1B. In Munc13-1, Glu-582 is exclusively photolabeled by the azialcohols. Azialcohols failed to photolabel these sites when these residues were mutated with alanine. Inspection of the model structure of PKCï¥C1 and Munc13-1C1 reveals that these residues forms groove where alcohol can bind. The present results provide evidence for the presence of alcohol-binding sites on PKCï¥ and Munc13-1 and underscore the power of mass spectrometry in identifying sites of alcohol action in the brain.