Increasing the fidelity of noncanonical amino acid incorporation in cell-free protein synthesis.

ABSTRACT:

Background

Cell-free protein synthesis provides a robust platform for co-translational incorporation of noncanonical amino acid (ncAA) into proteins to facilitate biological studies and biotechnological applications. Recently, eliminating the activity of release factor 1 has been shown to increase ncAA incorporation in response to amber codons. However, this approach could promote mis-incorporation of canonical amino acids by near cognate suppression.

Methods

We performed a facile protocol to remove near cognate tRNA isoacceptors of the amber codon from total tRNAs, and used the phosphoserine (Sep) incorporation system as validation. By manipulating codon usage of target genes and tRNA species introduced into the cell-free protein synthesis system, we increased the fidelity

SUBMITTER: Gan Q 

PROVIDER: S-EPMC5457342 | biostudies-literature | 2017 Nov

REPOSITORIES: biostudies-literature