A possible approach to site-specific insertion of two different unnatural amino acids into proteins in mammalian cells via nonsense suppression.
The site-specific insertion of an unnatural amino acid into proteins in vivo via nonsense suppression has resulted in major advances in recent years. The ability to incorporate two different unnatural amino acids in vivo would greatly increase the scope and impact of unnatural amino acid mutagenesis. Here, we show the concomitant suppression of an amber and an ochre codon in a single mRNA in mammalian cells by importing a mixture of aminoacylated amber and ochre suppressor tRNAs. This result provides a possible approach to site-specific insertion of two different unnatural amino acids into any protein of interest in mammalian cells. To our knowledge, this result also represents the only demonstration of concomitant suppression of two different termination codons in a single gene in vivo.
Köhrer
Caroline
C
Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
Yoo
Jae-Ho
JH
Bennett
Michael
M
Schaack
Jerome
J
RajBhandary
Uttam L
UL
D000596:Amino Acids / Q000737:chemistry*; D000818:Animals; D019556:COS Cells; D002460:Cell Line; D003062:Codon; D006801:Humans; D008156:Luciferases / Q000235:genetics; D011506:Proteins / Q000737:chemistry*; D012343:RNA, Transfer / Q000737:chemistry
D000596:Amino Acids; D003062:Codon; D011506:Proteins; D012343:RNA, Transfer; D008156:Luciferases