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Date of Award
Tamara L. Hendrickson
INVESTIGATING THE GLUTAMINE-TRNA (GLUTAMINE) SYNTHESIS APPARTUS OF THE HUMAN PATHOGEN HELICOBACTER PYLORI
Advisor: Tamara L. Hendrickson
Degree: Master of Science
Accurate protein biosynthesis is a vital process to all cellular life. Aminoacyl-tRNAs are at the heart of this process: A correctly formed aminoacyl-tRNA is critical for protein biosynthesis. Organisms have evolved many mechanisms to repair misacylated tRNAs before they cause errors in protein biosynthesis, thus maintaining the integrity of the genetic code. The human pathogen Helicobacter pylori (H. pylori) synthesizes Glu-tRNAGln as an intermediate to producing Gln-tRNAGln. This misacylated intermediate could cause lethal errors if used by the ribosome for protein synthesis. H. pylori repairs this intermediate by the amidotransferase GatCAB.
This dissertation focuses on indirect aminoacylation and transamidation to produce Gln-tRNAGln in H. pylori. A combination of phylogenetic analyses and enzymatic assays were used that contribute to a picture of this process.
Chapter 2 discusses our examination of sequence conservation of gltX2 (the gene that encodes GluRS2) across different H. pylori strains. The conclusions of this sequencing effort are compared with trends in sequences of genes important in the aminoacylation step of protein synthesis.
Chapter 3 discusses one aspect of Gln-tRNAGln production - the mechanism of transport of ammonia from the active site of one subunit (GatA) to the active site of another (GatB) through an intramolecular hydrophilic tunnel. Site-directed mutagenesis of key residues lining this tunnel and their transamidation assays were performed. With these studies, a preliminary picture of ammonia transport through the tunnel can be constructed.
Joshi, Nilesh, "Investigating the glutamine-trna (glutamine) synthesis appartus of the human pathogen helicobacter pylori" (2012). Wayne State University Theses. Paper 195.