Project Summary/Abstract Overview of Research in the Lorigan Lab and 5 Year Goals (Overview): Currently, we have limited structural information on membrane proteins. The Lorigan lab is interested in developing new biophysical methods to probe the structural and dynamic properties of integral membrane proteins using state-of-the-art pulsed EPR spectroscopic techniques and membrane solubilizing polymers. The overall objective is to study membrane proteins with EPR in a lipid bilayer as opposed to a detergent because it more closely mimics a cell membrane. Several proteins have been shown to not function or fold up correctly in a micelle when compared to a lipid bilayer. This is challenging because it is more difficult to express, purify, and conduct biophysical spectroscopic experiments on membrane proteins in lipid bilayers when compared to micelle or globular systems. My expertise in membrane protein EPR and sample preparation coupled with the powerful pulsed EPR instrumentation (DEER and ESEEM) that can measure long range distances has attracted several significant collaborators with important biological problems. My research lab works directly with several researchers to dramatically improve the quality of membrane protein sample preparation to yield high quality DEER data that leads to more accurate structural information. Please see the letters of support. The major biological focus of the lab is on investigating the structural and dynamic properties of integral membrane proteins in the KCNE family which is responsible for the modulation of voltage gated potassium (Kv) channels including KCNQ1 (Q1). This channel is expressed throughout the body to regulate physiological functions and mutations in this channel are linked to several diseases such as Long QT syndrome and other heart diseases. Increased understanding of the molecular underpinnings of KCNE modulation of potassium channels, specifically Q1, would help expand our understanding of the etiology of these diseases. We will study three members of the KCNE family: KCNE1(E1), KCNE3(E3), and KCNE4(E4). We are currently applying state-of-the-art EPR techniques to directly probe the structural and dynamic properties of Q1, E1, E3, and E4. This information will help us better understand Q1 regulation. The following pertinent biological questions will be answered: Which segments of E3 and E4 are helical in a lipid bilayer? What is the structure and topology of the KCNE3 and KCNE4 with respect to the membrane? How does E1 bind and interact with Q1 that is required for function? (5 Year Goals of the Lab): (1) Develop new biophysical techniques to dramatically improve structural studies of membrane proteins, (2) Probe the structural and dynamic properties of E3, and E4, (3) Elucidate the binding mechanism of E1 and Q1 to help us better understand the mechanism of Q1 regulation, and (4) Apply the membrane protein techniques that we develop to investigate the structure of several biologically impo...