Genetic Code Expansion of Mammalian Cells with Unnatural Amino Acids.
The expansion of the genetic code of mammalian cells enables the incorporation of unnatural amino acids into proteins. This is achieved by adding components to the protein biosynthetic machinery, specifically an engineered aminoacyl-tRNA synthetase/tRNA pair. The unnatural amino acids are chemically synthesized and supplemented to the growth medium. Using this methodology, fundamental new chemistries can be added to the functional repertoire of the genetic code of mammalian cells. This protocol outlines the steps necessary to incorporate a photocaged lysine into proteins and showcases its application in the optical triggering of protein translocation to the nucleus.
University of Pittsburgh, Department of Chemistry, Pittsburgh, Pennsylvania.
Kalyn A
KA
Brown
University of Pittsburgh, Department of Chemistry, Pittsburgh, Pennsylvania.
Alexander
A
Deiters
D000596:Amino Acids / Q000737:chemistry* / Q000378:metabolism; D000818:Animals; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D005815:Genetic Code; D008322:Mammals / Q000235:genetics* / Q000378:metabolism; D021381:Protein Transport; D011506:Proteins / Q000378:metabolism; D019680:Trinucleotide Repeat Expansion
D000596:Amino Acids; D011506:Proteins
caging; genetic code expansion; mammalian cell; nuclear localization; optogenetics; protein translocation; unnatural amino acid