Method Development Toward the Analysis of the Functionalization of Proteins

Calcote, Victoria (2016) Method Development Toward the Analysis of the Functionalization of Proteins. Undergraduate thesis, under the direction of Susan Pedigo from Chemistry & Biochemistry, The University of Mississippi.

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Calmodulin is a ubiquitous small intracellular protein that functions as a receptor for regulatory calcium signals and thus regulates a multitude of physiological processes in organisms as diverse as yeast, fruit flies, and mammals. Since its structure and function have been extensively studied, our goal is to create a calmodulin biomaterial, with the ultimate goal of in situ drug delivery or as a scaffold for stem cell growth and differentiation. We plan to create this biomaterial by using “Click Chemistry,” the rapid creation of new heteroatom bonds through a copper-catalyzed azide-alkyne cycloaddition. The bonds are created by treating calmodulin as an organic reagent and derivatizing the hydroxyl group of the amino acid tyrosine for an organic fluorophore. However, since tyrosines appear at residues 99 and 138, functionalization could be occurring at multiple sites. A reliable protocol has to be developed to determine the exact position of functionalization on calmodulin. We will focus on determining if and where functionalization occurs by employing an enzyme to cleave between the two tyrosines. Thrombin, a protease, offers a convenient analytical solution by cleaving the protein between two tyrosines, as it hydrolyzes the protein backbone at the C-terminal side of arginine (residue 106), which is located in between the tyrosines (residues 99 and 138). In finding a mechanism through which we are able to successfully separate the tyrosines on calmodulin, we will determine which, if either, has been derivatized by then performing gel electrophoresis, mass spectrometry, or chromatography.

Item Type: Thesis (Undergraduate)
Creators: Calcote, Victoria
Student's Degree Program(s): B.A. in Biochemistry
Thesis Advisor: Susan Pedigo
Thesis Advisor's Department: Chemistry & Biochemistry
Institution: The University of Mississippi
Subjects: Q Science > QD Chemistry
Depositing User: Victoria Calcote
Date Deposited: 13 May 2016 13:35
Last Modified: 13 May 2016 13:35

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