Membrane proteins are often used as targets for development of new drugs. Unfortunately, it is often difficult to determine the structures of these membrane proteins in their native environment due to hydrophobic interactions. It is estimated that 25% of all proteins are membrane proteins, yet structures have only been solved for a few hundred.
Holins are a class of proteins that form holes in bacterial membranes as part of the bacteriophage cell lysis mechanism. Pinholins form holes on the scale of 1.5 nm, compared to the 400 nm holes formed by Holin proteins. These holes allow the free flow of ions across the membrane, resulting in its depolarization and the activation of Signal Anchor-Release (SAR) endolysins. The S21 pinholin protein is encoded by the ϕ21 bacteriophage and contains two trans-membrane domains. Identification of its structural conformations will allow for better characterization of the lysis pathway: a possible target for antiviral therapy.
Our lab synthesizes and purifies the pinholin protein before incorporating it in a lipid bilayer for analysis by electron paramagnetic resonance (EPR). In this study, we used continuous wave (CW) EPR and double electron-electron repulsion (DEER) spectroscopy to obtain structural information on the pinholin protein. Specifically, we used CW-EPR to determine how much different parts of the protein structure move when the protein is incorporated in a bilayer, and DEER to measure the distance between two locations on the protein spin-labelled with MTSL. Using this information, we were able to model the native structure of the pinholin protein in a membrane mimetic environment.
Author: Jack Bennett
Faculty Advisor: Gary Lorigan, Department of Chemistry and Biochemistry
Graduate Student Advisors: Tanbir Ahammad, Department of Chemistry and Biochemistry, and Rasal Khan, Department of Chemistry and Biochemistry
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