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Journal Article

Prp2-mediated protein rearrangements at the catalytic core of the spliceosome as revealed by dcFCCS.

MPS-Authors
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Ohrt,  T.
Department of Cellular Biochemistry, MPI for biophysical chemistry, Max Planck Society;

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Dannenberg,  J.
Department of Cellular Biochemistry, MPI for biophysical chemistry, Max Planck Society;

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Odenwälder,  P.
Department of Cellular Biochemistry, MPI for biophysical chemistry, Max Planck Society;

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Dybkov,  O.
Department of Cellular Biochemistry, MPI for biophysical chemistry, Max Planck Society;

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Rasche,  N.
Department of Cellular Biochemistry, MPI for biophysical chemistry, Max Planck Society;

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Schmitzova,  J.
Department of Cellular Biochemistry, MPI for biophysical chemistry, Max Planck Society;

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Fabrizio,  P.
Department of Cellular Biochemistry, MPI for biophysical chemistry, Max Planck Society;

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Lührmann,  R.
Department of Cellular Biochemistry, MPI for biophysical chemistry, Max Planck Society;

External Ressource
Fulltext (public)

1481386.pdf
(Publisher version), 898KB

Supplementary Material (public)

1481386_Supplement.pdf
(Supplementary material), 2MB

Citation

Ohrt, T., Prior, M., Dannenberg, J., Odenwälder, P., Dybkov, O., Rasche, N., et al. (2012). Prp2-mediated protein rearrangements at the catalytic core of the spliceosome as revealed by dcFCCS. RNA, 18(6), 1244-1256. doi:10.1261/rna.033316.112.


Cite as: http://hdl.handle.net/11858/00-001M-0000-000F-A623-6
Abstract
The compositional and conformational changes during catalytic activation of the spliceosome promoted by the DEAH box ATPase Prp2 are only poorly understood. Here, we show by dual-color fluorescence cross-correlation spectroscopy (dcFCCS) that the binding affinity of several proteins is significantly changed during the Prp2-mediated transition of precatalytic Bact spliceosomes to catalytically activated B* spliceosomes from Saccharomyces cerevisiae. During this step, several proteins, including the zinc-finger protein Cwc24, are quantitatively displaced from the B* complex. Consistent with this, we show that Cwc24 is required for step 1 but not for catalysis per se. The U2-associated SF3a and SF3b proteins Prp11 and Cus1 remain bound to the B* spliceosome under near-physiological conditions, but their binding is reduced at high salt. Conversely, high-affinity binding sites are created for Yju2 and Cwc25 during catalytic activation, consistent with their requirement for step 1 catalysis. Our results suggest high cooperativity of multiple Prp2-mediated structural rearrangements at the spliceosome’s catalytic core. Moreover, dcFCCS represents a powerful tool ideally suited to study quantitatively spliceosomal protein dynamics in equilibrium.