Title

Screening Cellular Machinery for New Unnatural Amino Acids

Faculty Sponsor

Laura Rowe

College

Arts and Sciences

Department/Program

Chemistry

Document Type

Poster Presentation

Symposium Date

Summer 7-31-2014

Abstract

Orthogonal tRNA/aminoacyl tRNA synthetase pairs allow the incorporation of unnatural amino acids into specific locations within a protein structure in vivo. Current screening methods to determine whether or not tRNA/aaRS pairs incorporate a specific unnatural amino acid fail to de-select pairs that can incorporate multiple unnatural amino acids beyond the one for which they are designed. A recently developed method allows for this screening of promiscuous unnatural amino acid incorporation using tRNA/aaRS pairs in conjunction with a green fluorescent protein (GFP) with a stop codon mutation at a central, non-essential position. We are recreating this GFP based screening system in order to test for the incorporation of fluorescent unnatural amino acids into proteins using these promiscuous tRNA/aaRS pairs. Thus far, we have tested the fluorescence and solvatochromic properties of the fluorescent unnatural amino acids dansylalanine, p-cyanophenylalanine, and natural tryptophan. We also constructed the screening system with several tRNA/aaRS pair plasmids and one GFP plasmid mutated with a centrally located stop codon. We are currently testing these fluorescent unnatural amino acids (dansylalanine and p-cyanophenylalanine) in our designed GFP based screening system, in order to see if they are incorporated into protein via the poly-specificity of the tRNA/aaRS pairs.

Biographical Information about Author(s)

I am a senior biology and chemistry double major here at Valparaiso. I am hoping to use my research experience to help me prepare for dental school in terms of research preparedness through literature and critical thinking, as well as manual dexterity within lab. I became interested in the study of fluorescent unnatural amino acids during my biochemistry laboratory, and am glad I have had the chance to study them in more detail.

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