Ribosome selectivity and nascent chain context in modulating the incorporation of fluorescent non-canonical amino acid into proteins.

ABSTRACT: Fluorescence reporter groups are important tools to study the structure and dynamics of proteins. Genetic code reprogramming allows for cotranslational incorporation of non-canonical amino acids at any desired position. However, cotranslational incorporation of bulky fluorescence reporter groups is technically challenging and usually inefficient. Here we analyze the bottlenecks for the cotranslational incorporation of NBD-, BodipyFL- and Atto520-labeled Cys-tRNA^Cys into a model protein using a reconstituted in-vitro translation system. We show that the modified Cys-tRNA^Cys can be rejected during decoding due to the reduced ribosome selectivity for the modified aa-tRNA and the competition with native near-cognate aminoacyl-tRNAs. Accommodation of the modified Cys-tRNA<

SUBMITTER: Thommen M 

PROVIDER: S-EPMC9329280 | biostudies-literature | 2022 Jul

REPOSITORIES: biostudies-literature