Dissertation Defense Announcement
To:  The George Mason University Community

Candidate: Ekaterina Marakasova
Program: PhD in Biosciences
 
Date:   Monday May 21, 2018
Time:   12:00 noon
Place:  George Mason University
 Science & Tech Campus
  Institute for Advanced Biomedical Research, Room 1003  
 
Title: "Post-translational Modifications of Francisella tularensis Proteome”

Committee Chair: Dr. Monique van Hoek
Committee Members:  Dr. Ancha Baranova, Dr. Iosif Vaisman, Dr. Siobhán Cowley

All are invited to attend the defense.

ABSTRACT:
Post-translational modifications (PTMs) generally refer to covalent addition of a chemical group to the protein by an enzyme. There are more than 200 known PTMs that result in protein mass change, hydrophobicity and charge that influence protein folding and conformation and therefore allow cells rapidly respond to constantly changing environmental conditions. Francisella tularensis SCHU S4 is a gram-negative bacterium that causes human disease called tularemia, a disease that has been reported by CDC from all US states except Hawaii. Due to its high infectivity and ability to be transmitted by air, F. tularensis has been categorized as a class A bioterrorism agent. Francisella tularensis genome encodes a little less than 2000 proteins; however, PTMs are identified for a very small number of them. This dissertation is focused on two distinct PTMs of Francisella tularensis proteome: prenylation and N-ɛ-lysine acetylation.

Prenylation is a covalent addition of hydrophobic group to the C terminus of the protein, such modification is very important in protein folding, activity and cellular localization. We performed a global analysis of prenylation of pathogenic microorganisms by bioinformatics methods, suggesting that a number of pathogen-encoded proteins might be prenylated by host-cell machinery. Use of prenylation inhibitors as potential antimicrobial drugs is discussed.

N-acetylation is a reversible enzymatic PTM in which an acetyl group is added to the ɛ-amine group of the lysine. The lysine acetylation proteome profiling was done using enrichment of lysine-acetylated tryptic peptides/proteins with anti-acetyl-lysine antibodies and subsequent peptide identification by Mass Spectrometry (LC/MS/MS). Acetylation sites were validated with two acetylation prediction programs. We discovered 72 acetylated proteins with 135 sites of acetylation, which are involved in diverse metabolic processes such as the TCA cycle, carbon metabolism, fatty acid metabolism, transcription, translation, stress response and infectious process. The proteins were characterized by Gene Ontology annotations; the protein interaction network was built with further MCL clustering; the subcellular localization was determined by PSORT. The acetylation site was characterized by amino acid sequence properties and secondary structures. Such top-down approach allowed us to determine chitinase A (ChiA) is lysine acetylated, which is consistent with our bioinformatics predictions. ChiA acetylation in vitro can downregulate the activity of the enzyme, which suggests possible regulation mechanism of enzymatic activity by acetylation in Francisella tularensis.

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