PDBsum entry 2ghp

RNA binding protein

PDB id: 2ghp

Contents

Protein chains

(+ 2 more) 232 a.a. *

Waters ×193

* Residue conservation analysis

PDB id:

2ghp

Name:

RNA binding protein

Title:

Crystal structure of the n-terminal 3 RNA binding domains of the yeast splicing factor prp24

Structure:

U4/u6 snrna-associated splicing factor prp24. Chain: a, b, c, d, e, f, g, h. Synonym: u4/u6 snrnp protein. Engineered: yes

Source:

Saccharomyces cerevisiae. Baker's yeast. Organism_taxid: 4932. Gene: prp24. Expressed in: escherichia coli. Expression_system_taxid: 562.

Biol. unit:

Tetramer (from PQS)

Resolution:

2.70Å R-factor: 0.213 R-free: 0.264

Authors:

E.Bae,G.E.Wesenberg,G.N.Phillips Jr.,E.Bitto,C.A.Bingman,Center For Eukaryotic Structural Genomics (Cesg)

Key ref:

E.Bae et al. (2007). Structure and interactions of the first three RNA recognition motifs of splicing factor prp24. J Mol Biol, 367, 1447-1458. PubMed id: 17320109 DOI: 10.1016/j.jmb.2007.01.078

Date:

27-Mar-06 Release date: 25-Apr-06

Protein chains

P49960  (PRP24_YEAST) -  U4/U6 snRNA-associated-splicing factor PRP24 from Saccharomyces cerevisiae (strain ATCC 204508 / S288c)

Seq: 444 a.a.

Struc: 232 a.a.

DOI no: 10.1016/j.jmb.2007.01.078

J Mol Biol 367:1447-1458 (2007)

PubMed id: 17320109

Structure and interactions of the first three RNA recognition motifs of splicing factor prp24.

E.Bae, N.J.Reiter, C.A.Bingman, S.S.Kwan, D.Lee, G.N.Phillips, S.E.Butcher, D.A.Brow.

ABSTRACT

The essential Saccharomyces cerevisiae pre-messenger RNA splicing protein 24 (Prp24) has four RNA recognition motifs (RRMs) and facilitates U6 RNA base-pairing with U4 RNA during spliceosome assembly. Prp24 is a component of the free U6 small nuclear ribonucleoprotein particle (snRNP) but not the U4/U6 bi-snRNP, and so is thought to be displaced from U6 by U4/U6 base-pairing. The interaction partners of each of the four RRMs of Prp24 and how these interactions direct U4/U6 pairing are not known. Here we report the crystal structure of the first three RRMs and the solution structure of the first two RRMs of Prp24. Strikingly, RRM 2 forms extensive inter-domain contacts with RRMs 1 and 3. These contacts occupy much of the canonical RNA-binding faces (beta-sheets) of RRMs 1 and 2, but leave the beta-sheet of RRM 3 exposed. Previously identified substitutions in Prp24 that suppress mutations in U4 and U6 spliceosomal RNAs cluster primarily in the beta-sheet of RRM 3, but also in a conserved loop of RRM 2. RNA binding assays and chemical shift mapping indicate that a large basic patch evident on the surface of RRMs 1 and 2 is part of a high affinity U6 RNA binding site. Our results suggest that Prp24 binds free U6 RNA primarily with RRMs 1 and 2, which may remodel the U6 secondary structure. The beta-sheet of RRM 3 then influences U4/U6 pairing through interaction with an unidentified ligand.

Selected figure(s)

Figure 4.
Figure 4. Solution structure of Prp24-12. (a) Ensemble of ten energy-minimized NMR structures of Prp24-12. The backbone RMSD of secondary structure-containing regions of RRMs 1 (yellow) and 2 (orange) is 0.97(±0.09) Å. (b) Ribbon diagram of the lowest free energy structure of Prp24-12, with RRMs colored as in Figure 1, Figure 2 and Figure 3. (c) Comparison of the mean NMR and crystal (depicted in grey) structures. Backbone atoms of secondary structure regions were fit to RRM domains 1 and 2 and give an RMSD of 2.0(±0.3) Å (Supplementary Table III).

Figure 8.
Figure 8. Mapping of U6 RNA binding interactions on the surface of Prp24. (a) Region of the ^1H-^15N HSQC-TROSY spectra showing amide peaks of 0.2 mM ^2H,^15N-labeled Prp24-N12 protein in the absence (black) and presence (red) of 50 μM 21-nucleotide RNA (yeast U6 nucleotides 40–60). Assigned resonances that are selectively line broadened upon the addition of RNA are circled in magenta and labeled. (b) Location of the line broadened amino acid residues (shown in magenta) in the crystal structure. The structure at left is in the same orientation as is shown in Figure 3(a) and (b), and is rotated 180° around the vertical axis relative to the structure at right. The ribbon is colored as in Figure 1. (c) Electrostatic potential isosurface of the Prp24-N123 crystal structure calculated by the Poisson-Boltzmann equation. The isosurface is contoured at ±4 kT and is superposed on the molecular surface (white). A prominent electropositive region (blue) exists on one side of the protein, possibly corresponding to an RNA binding site. Electronegative regions (red) are small and distributed more uniformly around the protein. Structures are in the same orientation as in (b).

The above figures are reprinted by permission from Elsevier: J Mol Biol (2007, 367, 1447-1458) copyright 2007.

Figures were selected by the author.