Date of Award

12-2013

Degree Type

Thesis

Degree Name

Departmental Honors

Department

Chemistry and Biochemistry

Abstract

Many essential physiological pathways, such as cell proliferation, gene expression, and cardiovascular health are regulated by Protein Arginine Methyltransferases (PRMTs) through methylation of arginine residues in protein substrates. Understanding how PRMTs interact with their substrates is pivotal to understanding the biological role of these enzymes, and fundamental to the goal of identifying possible sites to be inhibited through drug therapy. Natural variations in the N-terminus of the PRMTl enzymes and data collected in our lab suggest that the N-terminus is important for activity and/or the binding of protein substrates. Preliminary data collected had led us to hypothesize that the negatively charged residues in the N-terminus (Nt) of PRMT1 play an important role in substrate specificity and protein-substrate interactions. This paper details the exploration of this hypothesis through the creation of DNA vectors for the E. coli expression of mutant human PRMT1 proteins in which varying negatively charged residues are mutated to non-charged residues. These vectors were transformed into E. coli cells and expression and purification of each mutant was partially optimized. Of the three hPRMT1 mutants created, one has been kinetically characterized and the activity of this mutant on three wild type PRMT1 substrates has been examined. The ultimate aim of this study, accompanied with ongoing studies in the Hevel laboratory, is to form a complete picture of how PRMT1 interacts with its substrates and the role of the N-terminus of PRMT1 in this process.

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Faculty Mentor

Joanie M. Hevel

Departmental Honors Advisor

Alvan Hengge