Abstract
An unnatural base pair between 7-(2-thienyl)-imidazo[4,5-b]pyridine (Ds) and pyrrole-2-carbaldehyde (Pa) could expand the genetic alphabet and allow the incorporation of non-standard amino acids into proteins at defined positions. For this purpose, we synthesized tRNAs bearing Pa at the anticodon and tested non-standard amino acid phosphoserine aminoacylation by the wild-type and various engineered phosphoseryl-tRNA synthetases (SepRSs). The D418N D420N T423V triple mutant of SepRS efficiently charged phosphoserine to tRNA containing the PaUA anticodon with a Km = 47.1 μM and a kcat = 0.151 s-1, which are comparable to the values of the wild-type SepRS for its cognate substrate, tRNACys with the GCA anticodon (26.9 μM and 0.111 s-1). The triple mutant SepRS and the tRNA with the PaUA anticodon represent a specific pair for the site-specific incorporation of phosphoserine into proteins in response to the UADs codon within mRNA.
Original language | English (US) |
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Pages (from-to) | 480-485 |
Number of pages | 6 |
Journal | Biochemical and Biophysical Research Communications |
Volume | 372 |
Issue number | 3 |
DOIs | |
State | Published - Aug 1 2008 |
Externally published | Yes |
Keywords
- Aminoacylation
- Engineering
- Phosphoserine
- Synthetase
- Unnatural base pair
- tRNA
ASJC Scopus subject areas
- Biophysics
- Biochemistry
- Molecular Biology
- Cell Biology