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Intern: Jaclyn Martinez Mentor: Luke Sherlin Faculty Supervisor: Professor John J. Perona Department :Chemistry and Biochemistry

PURIFICATION AND CRSTYALLIZATION OF GLN-tRNA^Gln PROTEIN COMPLEXES

During protein synthesis, the genetic code embedded in DNA is transcribed and translated to create a protein chain. Messenger RNA (mRNA) contains the genetic code in a series of trinucleotide units or codons. Transfer RNA (tRNA) provides a link between the mRNA sequence and the protein chain by carrying an amino acid that corresponds with the complementary codon of the tRNA. Some structures of tRNA are known but no structures of human tRNA have been determined. Through my research, the kinetic parameters of the human tRNA^Gln will be calculated from aminoacylation assays. The tRNA construct will then be crystallized, and x-ray crystallography will be used to determine the molecular structure. The synthesis and purification of tRNA constructs involves first a DNA polymerase, which is used to construct a DNA template from which human tRNA is translated using a Bacteriophage T7 RNA polymerase. Once the correct product is verified, the tRNA^Gln is purified through a variety of techniques and dialysized to ensure that the tRNA stays at a low salt concentration and at a constant pH. The techniques have been successful and produced a valid product. In order to evaluate the ability of E. coli glutaminyl-tRNA synthetase in acting on the human tRNA construct, aminoacylation assays will be run to determine the rate of aminoacylation. After which the tRNA will be crystallized for further study.

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