High Yield, Low Magnesium Flexizyme Reactions in a Water-Ice Eutectic Phase
Flexizymes enable the stoichiometric acylation of tRNAs with a variety of compounds, enabling the <i>in vitro</i> translation of peptides with both non-natural backbones and side chains. However, flexizyme reactions have several drawbacks, including single-turnover kinetics, high Mg(II) carryover, inhibiting <i>in vitro</i> translation, and rapid product hydrolysis. Here we present flexizyme reactions utilizing an ice-eutectic phase, with high yields, 30 times lower Mg(II), and long-term product stability. The eutectic flexizyme reactions increase the ease of use, yield and flexibility of aminoacylation and significantly increase the <i>in vitro</i> protein production.
Authors
- Isaac J. Knudson (ORCID: https://orcid.org/0000-0003-4773-0257)
- Mace Blank
- José Luís Pérez Alejo (ORCID: https://orcid.org/0000-0002-1952-4109)
- Evan M. Kalb
- Alanna Schepartz (ORCID: https://orcid.org/0000-0003-2127-3932)
- Aaron E. Engelhart (ORCID: https://orcid.org/0000-0002-1849-7700)
- Katarzyna P. Adamala (ORCID: https://orcid.org/0000-0003-1066-7207)
- Joshua A. Davisson (ORCID: https://orcid.org/0000-0001-9791-7816)
- Angelin Prasad
Institutions
- University of Minnesota (US)
- St. Jude Children's Research Hospital (US)
- University of Minnesota System (US)
- University of California System (US)
- GeneticaLens (United States) (US)
- Institut thématique Génétique, génomique et bioinformatique (FR)
- University of California, Berkeley (US)
Publication Details
- Journal
- Biochemistry
- Published
- 2025-08-20
- DOI
- https://doi.org/10.1021/acs.biochem.5c00371
- Citations
- 1
- Primary Topic
- Chemical Synthesis and Analysis
- Type
- article
- Field-Weighted Citation Impact
- 0.81
Funders
- National Science Foundation
- Howard Hughes Medical Institute
- Alfred P. Sloan Foundation