title: Ribosome Subunit Stapling for Orthogonal Translation in E. coli.
abstract: The creation of orthogonal large and small ribosomal subunits, which interact with each other but not with endogenous ribosomal subunits, would extend our capacity to create new functions in the ribosome by making the large subunit evolvable. To this end, we rationally designed a ribosomal RNA that covalently links the ribosome subunits via an RNA staple. The stapled ribosome is directed to an orthogonal mRNA, allowing the introduction of mutations into the large subunit that reduce orthogonal translation, but have minimal effects on cell growth. Our approach provides a promising route towards orthogonal subunit association, which may enable the evolution of key functional centers in the large subunit, including the peptidyl-transferase center, for unnatural polymer synthesis in cells.
authors.affiliation: Medical Research Council Laboratory of Molecular Biology, Francis Crick Avenue, Cambridge CB2 0QH (UK).
authors.forename: Stephen D
authors.initials: SD
authors.lastname: Fried
authors.affiliation: Medical Research Council Laboratory of Molecular Biology, Francis Crick Avenue, Cambridge CB2 0QH (UK).
authors.forename: Wolfgang H
authors.initials: WH
authors.lastname: Schmied
authors.affiliation: Medical Research Council Laboratory of Molecular Biology, Francis Crick Avenue, Cambridge CB2 0QH (UK).
authors.forename: Chayasith
authors.initials: C
authors.lastname: Uttamapinant
authors.affiliation: Medical Research Council Laboratory of Molecular Biology, Francis Crick Avenue, Cambridge CB2 0QH (UK). chin@mrc-lmb.cam.ac.uk.
authors.forename: Jason W
authors.initials: JW
authors.lastname: Chin
mesh_terms: D001483:Base Sequence; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics* / Q000254:growth & development; D005815:Genetic Code; D012333:RNA, Messenger / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D012335:RNA, Ribosomal / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D012270:Ribosomes / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
chemical_list: D012333:RNA, Messenger; D012335:RNA, Ribosomal
keywords: genetic code reprogramming; orthogonal ribosomes; ribosomal RNA engineering; synthetic biology; translation