A ribozyme that can charge a tRNA with amino acids and discriminate between cognate and non-cognate tRNAs is of interest because an RNA with this ability may have been a critical for translation in the transition from the RNA world. In addition, it could provide a tool for incorporating non-canonical amino acids for biotechnology applications. Here, we rationally engineer a ribozyme by fusing a tRNA binding module derived from a T-box riboswitch with a catalytic module (a flexizyme) to generate a ribozyme that can amino acylate a target tRNA. We demonstrate that this ribozyme be readily redesigned to alter tRNA specificity. This ribozyme is compatible with an in vitro translation system and could be used to recode a protein sequence to site-specifically incorporate a non-canonical amino acid.
The structure of such a polymerase ribozyme bound to RNA substrates comprising the template, primer, and nucleoside triphosphate (NTP) analog is presented, revealing how directed evolution shaped flanking elements around a highly conserved catalytic core derived from the ancestral class I ligase ribozyme.
Timothy S. Strutzenberg, David P. Horning, Wesley G. Cochrane et al.· bioRxiv· 0 citations
Abstract Aminoacyl-tRNA synthetases (AARSs) safeguard translational fidelity by coordinating amino acid activation and tRNA charging within distinct catalytic and editing domains. In leucyl-tRNA synthetase (LeuRS), the small, centrally located zinc-binding domain (ZN domain) sits at the crossroads of these functional c...
G. Hoffmann, Morana Dulic, I. Gruic‐Sovulj et al.· Nucleic Acids Research· 0 citations
Aminoacyl-tRNA synthetases (aaRSs) catalyze the attachment of amino acids (AAs) to their cognate tRNAs during protein synthesis. As aaRSs possess highly selective amino acid-binding sites, a distinct enzyme is generally required for each amino acid in the genetic code. Recently, genetic code expansion (GCE) has emerged...
Surendar R Jakka, Sandhya Jaiswal, K. M. Reddy et al.· Angewandte Chemie· 0 citations
Genetic code expansion with noncanonical amino acids (ncAAs) opens new opportunities for the design and engineering of proteins by broadening their chemical repertoire. Unfortunately, ncAA incorporation into proteins is limited both by a small collection of orthogonal aminoacyl-tRNA synthetases (aaRSs) and tRNAs and...
Kosuke Seki, Michael T. A. Nguyen, Petar I. Penev et al.· ACS Synthetic Biology· 0 citations
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