Abstract
Brr2 is a DExD/H-box helicase responsible for U4/U6 unwinding during spliceosomal activation. Brr2 contains two helicase-like domains, each of which is followed by a Sec63 domain with unknown function. We determined the crystal structure of the second Sec63 domain, which unexpectedly resembles domains 4 and 5 of DNA helicase Hel308. This, together with sequence similarities between Brr2's helicase-like domains and domains 1-3 of Hel308, led us to hypothesize that Brr2 contains two consecutive Hel308-like modules (Hel308-I and Hel308-II). Our structural model and mutagenesis data suggest that Brr2 shares a similar helicase mechanism with Hel308. We demonstrate that Hel308-II interacts with Prp8 and Snu114 in vitro and in vivo. We further find that the C-terminal region of Prp8 (Prp8-CTR) facilitates the binding of the Brr2-Prp8-CTR complex to U4/U6. Our results have important implications for the mechanism and regulation of Brr2's activity in splicing.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
MeSH terms
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Adenosine Triphosphatases / chemistry*
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Adenosine Triphosphatases / genetics
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Adenosine Triphosphatases / metabolism
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Animals
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Humans
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Molecular Sequence Data
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Protein Structure, Secondary*
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Protein Structure, Tertiary*
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RNA Helicases / chemistry*
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RNA Helicases / genetics
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RNA Helicases / metabolism
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Ribonucleoprotein, U4-U6 Small Nuclear / genetics
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Ribonucleoprotein, U4-U6 Small Nuclear / metabolism
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Ribonucleoprotein, U5 Small Nuclear / genetics
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Ribonucleoprotein, U5 Small Nuclear / metabolism
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Saccharomyces cerevisiae Proteins / chemistry*
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Saccharomyces cerevisiae Proteins / genetics
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Saccharomyces cerevisiae Proteins / metabolism
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Spliceosomes / chemistry*
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Spliceosomes / genetics
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Spliceosomes / metabolism
Substances
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PRP8 protein, S cerevisiae
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Ribonucleoprotein, U4-U6 Small Nuclear
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Ribonucleoprotein, U5 Small Nuclear
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SNU114 protein, S cerevisiae
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Saccharomyces cerevisiae Proteins
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Adenosine Triphosphatases
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BRR2 protein, S cerevisiae
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RNA Helicases