Date of Award
5-28-2009
Degree Type
Dissertation
Degree Name
Doctor of Philosophy (PhD)
Department
Chemistry
First Advisor
Markus W. Germann - Chair
Second Advisor
Alfons L. Baumstark
Third Advisor
Jerry C. Smith
Fourth Advisor
Kathryn B. Grant
Abstract
Potassium ion channels play a key role in the generation and propagation of action potentials. The S4-S5 linker peptide (L45) is believed to be responsible for the anesthetic/alcohol response of voltage-gated K+ channels. We investigated this region to define the structural basis of 1-alkanol binding site in dShaw2 K+ channel. L45 peptides derived from dShaw2 and hKv3.4 K+ channel, which, if part of the complete channel, demonstrate different sensitivity to 1-alcohols. Specifically, dShaw2 is alcohol sensitive and hKv3.4 is alcohol resistant. Structural analysis of L45 with NMR and CD suggested a direct correlation between alpha-helicity and the inhibition of dShaw2 channel by 1-butanol. We used CD and NMR to determine the structure of L45 peptides in micelles and vesicles. We measured spin-lattice relaxation time (T1) and determined the location and surface accessibility of L45 in micelles. These experiments confirm that L45 of dShaw2 adopts an α-helical conformation, partially buried in the membrane and parallel to the surface. The binding and accumulation of rev proteins to an internal loop of RRE (rev responsive element) of unspliced mRNA precursors is a key step of propagation of human immunodeficiency (HIV) virus. Molecules that interfere with this process can be expected to show anti-HIV activity. Our work is based on an assumption that zinc fingers could compete with rev proteins, therefore impeding the life cycle of HIV and stopping its infection. We studied the influence of different cations, anions, and the concentration of salts and osmolytes on the binding affinity with Polyacrylamide Gel Electrophoresis (PAGE) and Isothermal Titration Calorimetry (ITC). We conclude that the types of anions and/or cations and their concentrations affect the enthalpy and entropy of the binding interacitons. Using a gel assay, we confirm that there are three products in RNA-Protein reaction, and both EDTA and salts (and their concentrations) in the gel or samples interfere with RNA-protein complex mobility.
DOI
https://doi.org/10.57709/1059276
Recommended Citation
Qu, Xiaoguang, "NMR Study of Structure and Orientation of S4-S5 Linker Peptides from Shaw Related Potassium Ion Channels in Micelles and Binding of ZNF29R Protein to HIV RREIIBTR RNA." Dissertation, Georgia State University, 2009.
doi: https://doi.org/10.57709/1059276