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Synthesis and Incorporation of Unnatural Amino Acids To Probe and Optimize Protein Bioconjugations
Maza, Johnathan C. McKenna, Jaclyn R. Raliski, Benjamin K. Freedman, Matthew T. Young, Douglas D.
Maza, Johnathan C.
McKenna, Jaclyn R.
Raliski, Benjamin K.
Freedman, Matthew T.
Young, Douglas D.
Abstract
The utilization of unnatural amino acids (UAAs) in bioconjugations is ideal due to their ability to confer a degree of bioorthogonality and specificity. In order to elucidate optimal conditions for the preparation of bioconjugates with UAAs, we synthesized 9 UAAs with variable methylene tethers (2-4) and either an azide, alkyne, or halide functional group. All 9 UAAs were then incorporated into green fluorescent protein (GFP) using a promiscuous aminoacyl-tRNA synthetase. The different bioconjugations were then analyzed for optimal tether length via reaction with either a fluorophore or a derivatized resin. Interestingly, the optimal tether length was found to be dependent on the type of reaction. Overall, these findings provide a better understanding of various parameters that can be optimized for the efficient preparation of bioconjugates.
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2015-01-01
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Chemistry
DOI
10.1021/acs.bioconjchem.5b00424