New in Vitro Assay Measuring Direct Interaction of Nonsense Suppressors with the Eukaryotic Protein Synthesis Machinery
Authors
Ng, Martin YZhang, Haibo
Weil, Amy
Singh, Vijay
Jamiolkowski, Ryan
Baradaran-Heravi, Alireza
Roberge, Michel
Jacobson, Allan
Friesen, Westley
Welch, Ellen
Goldman, Yale E
Cooperman, Barry S
UMass Chan Affiliations
Microbiology and Physiological SystemsDocument Type
Journal ArticlePublication Date
2018-11-21Keywords
Premature termination codon (PTC)readthrough
ribosome
in vitro assay
eukaryotic translation
nonsense suppressors
Metadata
Show full item recordAbstract
Nonsense suppressors (NonSups) induce "readthrough", i.e., the selection of near cognate tRNAs at premature termination codons and insertion of the corresponding amino acid into nascent polypeptide. Prior readthrough measurements utilized contexts in which NonSups can promote readthrough directly, by binding to one or more of the components of the protein synthesis machinery, or indirectly, by several other mechanisms. Here we utilize a new, highly purified in vitro assay to measure exclusively direct nonsense suppressor-induced readthrough. Of 16 NonSups tested, 12 display direct readthrough, with results suggesting that such NonSups act by at least two different mechanisms. In preliminary work we demonstrate the potential of single molecule fluorescence energy transfer measurements to elucidate mechanisms of NonSup-induced direct readthrough, which will aid efforts to identify NonSups having improved clinical efficacy.Source
Ng MY, Zhang H, Weil A, Singh V, Jamiolkowski R, Baradaran-Heravi A, Roberge M, Jacobson A, Friesen W, Welch E, Goldman YE, Cooperman BS. New in Vitro Assay Measuring Direct Interaction of Nonsense Suppressors with the Eukaryotic Protein Synthesis Machinery. ACS Med Chem Lett. 2018 Nov 21;9(12):1285-1291. doi: 10.1021/acsmedchemlett.8b00472. PMID: 30613341; PMCID: PMC6295867.DOI
10.1021/acsmedchemlett.8b00472Permanent Link to this Item
http://hdl.handle.net/20.500.14038/51739PubMed ID
30613341Related Resources
This article is based on a previously available preprint in bioRxiv, https://doi.org/10.1101/330506.ae974a485f413a2113503eed53cd6c53
10.1021/acsmedchemlett.8b00472