UMMS Affiliation

RNA Therapeutics Institute; Graduate School of Biomedical Sciences

Publication Date

2020-08-14

Document Type

Article Postprint

Disciplines

Amino Acids, Peptides, and Proteins | Biochemistry, Biophysics, and Structural Biology | Genetics and Genomics

Abstract

Termination of protein biosynthesis is an essential step of gene expression, during which a complete functional protein is released from the ribosome. Premature or inefficient termination results in truncated, non-functional or toxic proteins that may cause disease. Indeed, more than 10% of human genetic diseases are caused by nonsense mutations leading to premature termination. Efficient and sensitive approaches are required to study eukaryotic termination mechanisms and to identify potential therapeutics that modulate termination. Canonical radioactivity-based termination assays are complex, report on a short peptide release, and are incompatible with high-throughput screening. Here we describe a robust and simple in vitro assay to study the kinetics of full-protein release. The assay monitors luminescence upon release of nanoluciferase from a mammalian pre-termination complex. The assay can be used to record time-progress curves of protein release in a high-throughput format, making it optimal for studying release kinetics and for high-throughput screening for small molecules that modulate the efficiency of termination.

Keywords

luminescence, protein release, translation termination

Rights and Permissions

This article, published in RNA, is available under a Creative Commons License (Attribution-NonCommercial 4.0 International), as described at http://creativecommons.org/licenses/by-nc/4.0.

DOI of Published Version

10.1261/rna.076588.120

Source

Susorov D, Egri S, Korostelev AA. Termi-Luc: a versatile assay to monitor full-protein release from ribosomes. RNA. 2020 Aug 14:rna.076588.120. doi: 10.1261/rna.076588.120. Epub ahead of print. PMID: 32817446. Link to article on publisher's site

Journal/Book/Conference Title

RNA (New York, N.Y.)

Related Resources

Link to Article in PubMed

PubMed ID

32817446

Creative Commons License

Creative Commons Attribution-Noncommercial 4.0 License
This work is licensed under a Creative Commons Attribution-Noncommercial 4.0 License

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