Orthogonal translation enables heterologous ribosome engineering in E. coli
Artikel i vetenskaplig tidskrift, 2021

The ribosome represents a promising avenue for synthetic biology, but its complexity and essentiality have hindered significant engineering efforts. Heterologous ribosomes, comprising rRNAs and r-proteins derived from different microorganisms, may offer opportunities for novel translational functions. Such heterologous ribosomes have previously been evaluated in E. coli via complementation of a genomic ribosome deficiency, but this method fails to guide the engineering of refractory ribosomes. Here, we implement orthogonal ribosome binding site (RBS):antiRBS pairs, in which engineered ribosomes are directed to researcher-defined transcripts, to inform requirements for heterologous ribosome functionality. We discover that optimized rRNA processing and supplementation with cognate r-proteins enhances heterologous ribosome function for rRNAs derived from organisms with ≥76.1% 16S rRNA identity to E. coli. Additionally, some heterologous ribosomes undergo reduced subunit exchange with E. coli-derived subunits. Cumulatively, this work provides a general framework for heterologous ribosome engineering in living cells.

Författare

Natalie S. Kolber

Broad Institute

Stanford University

Ranan Fattal

Broad Institute

Sinisa Bratulic

Broad Institute

Chalmers, Biologi och bioteknik, Systembiologi

Gavriela D. Carver

Broad Institute

Ahmed H. Badran

Broad Institute

Nature Communications

2041-1723 (ISSN)

Vol. 12 1 599

Ämneskategorier

Biokemi och molekylärbiologi

Medicinsk bioteknologi (med inriktning mot cellbiologi (inklusive stamcellsbiologi), molekylärbiologi, mikrobiologi, biokemi eller biofarmaci)

Annan industriell bioteknik

DOI

10.1038/s41467-020-20759-z

PubMed

33500394

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Senast uppdaterat

2021-02-11