Year of Award

2022

Document Type

Thesis

Degree Type

Master of Science (MS)

Degree Name

Chemistry (Analytical/Environmental Option)

Department or School/College

Chemistry and Biochemistry

Committee Chair

Bruce Bowler

Commitee Members

Christopher P. Palmer, Bruce Bowler, Nigel Priestley, J. Stephen Lodmell

Keywords

Capillary Electrophoresis, Nanodisc

Publisher

University of Montana

Subject Categories

Analytical Chemistry

Abstract

Guanine nucleotide-binding proteins, or G-proteins, are crucial to cell signaling by acting as molecular switches through interactions with membrane-bound receptors. G-proteins have become of greater interest due to the possibility of novel drug targets. The G-protein alpha subunit, Gα, binds GTP before localizing to the membrane to interact with other G-protein subunits and G-protein-coupled receptors (GPCRs). Gi lacks a hydrophobic region to promote localization to the membrane, but a co-translational lipidation of the N-terminal residue stabilizes this interaction.

Protein interactions with biological membranes are challenging to study due to the complex structure of membranes and the often-limited quantities of the sample protein. To simplify the system and structure of membranes, membrane mimics, such as nanodiscs, are often utilized. Capillary electrophoresis has been a proven technique for measuring protein interactions due to the small amounts of required sample. In this study, the interaction of Gα with styrene-maleic acid co-polymer belt nanodiscs was measured using nanodisc affinity capillary electrophoresis (NACE). The data obtained suggests a complicated kinetic system that might be dependent upon a third component or a conformation change of the protein. Here we show the potential for NACE to estimate dissociation constants for proteins with membranes and discuss the limitations of the technique.

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© Copyright 2022 Bethany R. Emeigh