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Structural insights into how Prp5 proofreads the pre-mRNA branch site

Nature. 2021-08; 
Zhenwei Zhang, Norbert Rigo, Olexandr Dybkov, Jean-Baptiste Fourmann, Cindy L Will, Vinay Kumar, Henning Urlaub, Holger Stark, Reinhard Lührmann
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Custom DNA/RNA Oligos … The template for in vitro transcription of the U257A Act pre-mRNA was purchased from Genscript. Pre-A complexes were assembled in a 252 ml splicing reaction containing 1.7 nM pre-mRNA; before preparation of cryo-EM grids, three peak gradient fractions were buffer … Get A Quote

摘要

During the splicing of introns from precursor messenger RNAs (pre-mRNAs), the U2 small nuclear ribonucleoprotein (snRNP) must undergo stable integration into the spliceosomal A complex-a poorly understood, multistep process that is facilitated by the DEAD-box helicase Prp5 (refs. ). During this process, the U2 small nuclear RNA (snRNA) forms an RNA duplex with the pre-mRNA branch site (the U2-BS helix), which is proofread by Prp5 at this stage through an unclear mechanism. Here, by deleting the branch-site adenosine (BS-A) or mutating the branch-site sequence of an actin pre-mRNA, we stall the assembly of spliceosomes in extracts from the yeast Saccharomyces cerevisiae directly before the A complex is formed. W... More

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