Eric Kool, Postdoctoral Faculty Sponsor
Control of RNA with quinone methide reversible acylating reagents.
Organic & biomolecular chemistry
Caging RNA by polyacylation (cloaking) has been developed recently as a simple and rapid method to control the function of RNAs. Previous approaches for chemical reversal of acylation (uncloaking) made use of azide reduction followed by amine cyclization, requiring 2-4 h for the completion of cyclization. In new studies aimed at improving reversal rates and yields, we have designed novel acylating reagents that utilize quinone methide (QM) elimination for reversal. The QM de-acylation reactions were tested with two bioorthogonally cleavable motifs, azide and vinyl ether, and their acylation and reversal efficiencies were assessed with NMR and mass spectrometry on model small-molecule substrates as well as on RNAs. Successful reversal both with phosphines and strained alkenes was documented. Among the compounds tested, the azido-QM compound A-3 displayed excellent de-acylation efficiency, with t1/2 for de-acylation of less than an hour using a phosphine trigger. To test its function in RNA caging, A-3 was successfully applied to control EGFP mRNA translation in vitro and in HeLa cells. We expect that this molecular caging strategy can serve as a valuable tool for biological investigation and control of RNAs both in vitro and in cells.
View details for DOI 10.1039/d1ob01713f
View details for PubMedID 34528657
Single nucleotide translation without ribosomes
2021; 13 (8): 751-+
The translation of messenger RNA sequences into polypeptide sequences according to the genetic code is central to life. How this process, which relies on the ribosomal machinery, arose from much simpler precursors is unclear. Here, we demonstrate that single nucleotides charged with an amino acid couple with amino acids linked to the 5'-terminus of an RNA primer in reactions directed by the nucleotides of an RNA template in dilute aqueous solution at 0 °C. When a mixture of U-Val, A-Gly and G-Leu competed for coupling to Gly-RNA, base pairing dictated which dipeptide sequence formed preferentially. The resulting doubly anchored dipeptides can retain their link to the primer for further extension or can be fully released under mild acidic conditions. These results show that a single-nucleotide-based form of translation exists that requires no more than oligoribonucleotides and anchored amino acids.
View details for DOI 10.1038/s41557-021-00749-4
View details for Web of Science ID 000678413700003
View details for PubMedID 34312504