Abstract
The incorporation of noncanonical amino acids has given protein chemists access to an expanded repertoire of amino acids. This methodology has significantly broadened the scope of protein engineering allowing introduction of amino acids with non-native functionalities, such as bioorthogonal reactive handles (azides and alkynes) and hydrophobic fluorinated side chains. Here, we describe the efficient residue-specific replacement of methionine by azidonorleucine in an engineered green fluorescent protein using a bacterial expression system to introduce a single reactive site for the strain-promoted azide-alkyne cycloaddition.
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Acknowledgment
This work was financially supported by Netherlands Organisation for Scientific Research (Rubicon grant 680-50-1407 and Gravitation program 024.001.035). The plasmids encoding the mutant MetRS and GFP were a kind gift from Prof. D. A. Tirrell (California Institute of Technology).
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van Eldijk, M.B., van Hest, J.C.M. (2018). Residue-Specific Incorporation of Noncanonical Amino Acids for Protein Engineering. In: Lemke, E. (eds) Noncanonical Amino Acids. Methods in Molecular Biology, vol 1728. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7574-7_8
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DOI: https://doi.org/10.1007/978-1-4939-7574-7_8
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