An easy tool to monitor the elemental steps of in vitro translation via gel electrophoresis of fluorescently labelled small peptides

RNA. 2023 Dec 26:rna.079766.123. doi: 10.1261/rna.079766.123. Online ahead of print.ABSTRACTSeveral methods are available to visualize and assess the kinetics and efficiency of elemental steps of protein biosynthesis. However, each of these methods has its own limitations. Here, we present a novel, simple and convenient tool for monitoring stepwise in vitro translation initiated by BODIPY-Met-tRNA. Synthesis and release of very short, 1 to 7 amino acids, BODIPY-labelled peptides, can be monitored using urea-polyacrylamide gel electrophoresis. Very short BODIPY-labelled oligopeptides might be resolved this way, in contrast to widely used Tris-tricine gel electrophoresis, which is suitable to separate peptides larger than 1 kDa. The method described in this manuscript allows one to monitor the steps of translation initiation, peptide transfer, translocation, and termination as well as their inhibition at an unprecedented single amino acid resolution.PMID:38164606 | DOI:10.1261/rna.079766.123

Source: RNA - January 2, 2024 Category: Genetics & Stem Cells Authors: Valeriya I Marina Medina Bidzhieva Andrey G Tereshchenkov Dmitry Orekhov Vladislava E Sagitova Nataliya V Sumbatyan Vadim N Tashlitsky Artem S Ferberg Tinashe P Maviza Pavel Kasatsky Olga Tolicheva Alena Paleskava Vladimir I Polshakov Ilya A Osterman Olga Source Type: research