Intrinsically Disordered Protein Ntr2 Modulates the Spliceosomal RNA Helicase Brr2

Biophysical Journal
Jan WollenhauptMarkus C Wahl

Abstract

Precursor messenger RNA splicing is mediated by the spliceosome, a large and dynamic molecular machine composed of five small nuclear RNAs and numerous proteins. Many spliceosomal proteins are predicted to be intrinsically disordered or contain large disordered regions, but experimental validation of these predictions is scarce, and the precise functions of these proteins are often unclear. Here, we show via circular dichroism spectroscopy, dynamic light scattering, and NMR spectroscopy that the yeast spliceosomal disassembly factor Ntr2 is largely intrinsically disordered. Peptide SPOT analyses, analytical size-exclusion chromatography, and surface plasmon resonance measurements revealed that Ntr2 uses an N-terminal region to bind the C-terminal helicase unit of the Brr2 RNA helicase, an enzyme involved in spliceosome activation and implicated in splicing catalysis and spliceosome disassembly. NMR analyses suggested that Ntr2 does not adopt a tertiary structure and likely remains disordered upon complex formation. RNA binding and unwinding studies showed that Ntr2 downregulates Brr2 helicase activity in vitro by modulating the fraction of helicase molecules productively bound to the RNA substrate. Our data clarify the nature o...Continue Reading

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Citations

Jul 5, 2019·Biochimica Et Biophysica Acta. Gene Regulatory Mechanisms·Javier Oroz, Douglas V Laurents
Feb 15, 2021·Biological Chemistry·Pascal Donsbach, Dagmar Klostermeier

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