Engineered ribosomes with tethered subunits for expanding biological function

Erik D. Carlson, Anne E. d’Aquino, Do Soon Kim, Emily M. Fulk, Kim Hoang, Teresa Szal, Alexander S. Mankin and Michael C. Jewett ()
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Erik D. Carlson: Northwestern University
Anne E. d’Aquino: Northwestern University
Do Soon Kim: Northwestern University
Emily M. Fulk: Northwestern University
Kim Hoang: Northwestern University
Teresa Szal: University of Illinois at Chicago
Alexander S. Mankin: University of Illinois at Chicago
Michael C. Jewett: Northwestern University

Nature Communications, 2019, vol. 10, issue 1, 1-13

Abstract: Abstract Ribo-T is a ribosome with covalently tethered subunits where core 16S and 23S ribosomal RNAs form a single chimeric molecule. Ribo-T makes possible a functionally orthogonal ribosome–mRNA system in cells. Unfortunately, use of Ribo-T has been limited because of low activity of its original version. Here, to overcome this limitation, we use an evolutionary approach to select new tether designs that are capable of supporting faster cell growth and increased protein expression. Further, we evolve new orthogonal Ribo-T/mRNA pairs that function in parallel with, but independent of, natural ribosomes and mRNAs, increasing the efficiency of orthogonal protein expression. The Ribo-T with optimized designs is able to synthesize a diverse set of proteins, and can also incorporate multiple non-canonical amino acids into synthesized polypeptides. The enhanced Ribo-T designs should be useful for exploring poorly understood functions of the ribosome and engineering ribosomes with altered catalytic properties.

Date: 2019
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DOI: 10.1038/s41467-019-11427-y

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