An orthogonalized platform for genetic code expansion in both bacteria and eukaryotes.

Abstract:

:In this study, we demonstrate the feasibility of expanding the genetic code of Escherichia coli using its own tryptophanyl-tRNA synthetase and tRNA (TrpRS-tRNATrp) pair. This was made possible by first functionally replacing this endogenous pair with an E. coli-optimized counterpart from Saccharomyces cerevisiae, and then reintroducing the liberated E. coli TrpRS-tRNATrp pair into the resulting strain as a nonsense suppressor, which was then followed by its directed evolution to genetically encode several new unnatural amino acids (UAAs). These engineered TrpRS-tRNATrp variants were also able to drive efficient UAA mutagenesis in mammalian cells. Since bacteria-derived aminoacyl-tRNA synthetase (aaRS)-tRNA pairs are typically orthogonal in eukaryotes, our work provides a general strategy to develop additional aaRS-tRNA pairs that can be used for UAA mutagenesis of proteins expressed in both E. coli and eukaryotes.

journal_name

Nat Chem Biol

journal_title

Nature chemical biology

authors

Italia JS,Addy PS,Wrobel CJ,Crawford LA,Lajoie MJ,Zheng Y,Chatterjee A

doi

10.1038/nchembio.2312

subject

Has Abstract

pub_date

2017-04-01 00:00:00

pages

446-450

issue

4

eissn

1552-4450

issn

1552-4469

pii

nchembio.2312

journal_volume

13

pub_type

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