کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
2048490 | 1074082 | 2012 | 5 صفحه PDF | دانلود رایگان |

Posttranslational modifications play a crucial role in modulating protein structure and function. Genetic incorporation of unnatural amino acids into a specific site of a protein facilitates the systematic study of protein modifications including acetylation. We here report the directed evolution of pyrrolysyl-tRNA synthetase (PylRS) from Methanosarcina mazei to create N-acetyl lysyl-tRNA synthetases (AcKRSs) using a new selection system based on the killing activity of the toxic ccdB gene product. The amino acid specificity of these and of published [1] and [2] AckRSs was tested in vitro and in vivo, and the enzyme-kinetic properties of the AckRSs were evaluated for the first time.
► N-acetyl lysyl-tRNA synthetases were evolved by a CcdB-based selection.
► N-acetyl lysine specificity was validated by both in vitro and in vivo approaches.
► Kinetic properties of the evolved synthetases were evaluated for the first time.
► The evolved synthetases will facilitate the systematic study of protein acetylation.
Journal: FEBS Letters - Volume 586, Issue 6, 23 March 2012, Pages 729–733