Access Type

Open Access Thesis

Date of Award

January 2013

Degree Type

Thesis

Degree Name

M.S.

Department

Chemistry

First Advisor

David Rueda

Second Advisor

Louis J. Romano

Abstract

Splicing plays a major role in eukaryotic gene expression by processing pre-mRNA to form mature mRNA. Pre-mRNAs undergo splicing to remove introns, non–protein coding regions, and religate exons, protein coding regions. This process is catalyzed by the spliceosome, which consists of five small nuclear ribonucleoprotein particles (snRNPs: U1, U2, U4, U5 and U6) and numerous protein factors. Proper assembly of spliceosomal components is critical for function, and thus, defects in assembly can be lethal. Several spliceosomal proteins facilitate structural rearrangements important for spliceosomal assembly and function. Prp24 is an essential factor in U6 snRNP assembly, and it has been proposed to assist in U4/U6 formation and unwinding. Here, we address the question whether Prp24 affects the U2/U6 complex dynamics. Using single-molecule Fluorescence Resonance Energy Transfer (smFRET), we have previously shown that a minimal U2/U6 complex from yeast can adopt at least three distinct conformations in dynamic equilibrium. Our new single molecule data show that Prp24 unwinds U2 from U2/U6 complex and stabilizes U6 in a low FRET conformation. We also show that the RNA Recognition Motifs of Prp24 affect the binding affinity of Prp24 for U6 and unwinding activity. We propose that Prp24 plays an important role in U2 and U6 snRNP recycling by dissociating the U2/U6 complex.

Download

Share

COinS