Structural insight into RNA recognition motifs: versatile molecular Lego building blocks for biological systems

Schematic representation of the types of amino-acid residues on the β-sheet surfaces of RNA recognition motifs (RRMs). (a) Proportions of various amino-acid residues on the β-sheet surfaces of RRMs. The amino-acid residues are shown for (b) TIA-1 RRM2, (c) hTra2-β, (d) p14, (e) Nup35, and (f) Prp24 RRM4. The aromatic amino-acid residues and the Gly residue are colored red and cyan, respectively. The amino-acid residues with side chains directed toward the inside of the RRM fold are colored light gray, and those with solvent-exposed side chains are depicted in clear characters. In p14, Nup35, and Prp24 RRM4, the positions of the interacting amino-acid residues are shown with the three-letter code in a circle. The structures of the dimerized Nup35 RRM (PDB-ID: 1WWH) and the Prp24 RRM4 (PDB-ID: 2L9W) are demonstrated in the bottom panels of (e) and (f). These figures were generated by the program MOLMOL.21 (g) The RRM structures are represented by one block for a single RRM (top) and two blocks for tandem RRMs (bottom). The well-conserved β-sheet surface corresponds to the top surface of the box. Each of the surfaces is numbered from 1 to 6. In addition, the N- and C-terminal extensions are colored cyan and pink, respectively. In the case of the tandem RRMs, the surface numbers of the second RRM are underlined, and L indicates the linker between RRM1 and RRM2.

Involvement of the N- and/or C-terminal extension of the RNA recognition motif (RRM) in RNA recognition. (a) Structure of the U1A RRM1 in complex with stem II of the U1 snRNA (PDB-ID: 1URN). (b) Structure of the CUG-BP RRM3 in complex with 5′-UGUGUG-3′ (PDB-ID: 2RQC). (c) Structure of the PTB RRM3 in complex with 5′-UCUCU-3′ (PDB-ID: 2ADC). (d) Structure of the PTB RRM1 in complex with 5′-CUCU-3′ (PDB-ID: 2AD9). (e) Structure of the hTra2-β RRM in complex with 5′-GAAGAA-3′ (PDB-ID: 2RRA). (f) Structure of the C-terminal RRM of the U11/U12-65k protein (PDB-ID: 3EGN). The β-strands and α-helices of the RRM body are colored cyan and orange, respectively. The N-terminal extension that adopts the consecutive helical structure and holds the α2-helix of the RRM body is colored pink. The U11/U12-65k RRM contains an additional helical structure between the canonical α1–β2 loop. These figures were generated by the program MOLMOL.21

The appearance of the DxxT loop in RNA recognition motifs (RRMs). A total of 363 RRMs were aligned according to their homology to the TIA-1 RRM2. (a) The RRMs containing the DxxT loop sequence are demonstrated by a red bar. If the appearance of the DxxT loops correlated well with the total sequence homology, then the DxxT loops were clustered in the high homology region, as seen in the assumptive data. (b) However, the actual data indicated that the appearance of the DxxT loop is discretely identified.

RNA recognition on surfaces other than the β-sheet. (a) Poly(A)-specific ribonuclease RNA recognition motifs (PARN RRM) in complex with the m⁷GpppG cap structure (PDB-ID: 2ROK). (b) CBP20 in complex with the m⁷GpppG cap structure (PDB-ID: 1H2T). (c) The MTHFSD RRM (PDB-ID: 2E5J). (d) The hnRNP F RRM1 in complex with 5′-AGGGAU-3′ (PDB-ID: 2KFY). In (a), (b), and (d), the aromatic amino-acid residues that are involved in RNA recognition are colored orange, and those that interact with the N- and C-terminal extensions are colored red. In addition, in (d), the charged amino-acid residues involved in RNA binding are colored light green. In (c), a Trp residue is located at the position corresponding to the cap-interacting Trp residue of the PARN RRM. However, this Trp residue is involved in core formation in the MTHFSD RRM, and is buried. These figures were generated by the program MOLMOL.21

Cooperativity between multi RRMs. (A) The Sxl RRM1–RRM2 in complex with 5′-UGUUUUUUUU-3′ (PDB-ID: 1B7F). (b) The Hrp1 RRM1–RRM2 in complex with 5′-UAUAUAU-3′ (PDB-ID: 2CJK). (c) The PABP RRM1–RRM2 in complex with 5′-AAAAAAAA-3′ (PDB-ID: 1CVJ). (d) The PTB RRM3–RRM4 in complex with PPT (poly-pyrimidine tract) (PDB-ID: 2EVZ). (e) Hrp1/Rna15 in complex with 5′-UAUAUAUAAUAAU-3′ (PDB-ID: 2KM8). In (a) and (b), the key amino-acid residues that stack with the starting nucleotide on RRM1 are colored magenta. In (e), the inset indicates an expanded region, demonstrating the putative electrostatic interactions between the Rna15 RRM and the Hrp1 RRM1. These figures were generated by the program MOLMOL.21

Comparison between HuD and CUG-BP1. (a) HuD and CUG-BP1 share a similar domain architecture, with two consecutive N-terminal RRMs and a single C-terminal RRM. The complex structure of HuD RRM1–RRM2 with its target RNA (PDB-ID: 2FXL) and that of CUG-BP1 RRM3 are shown. In addition, the complex structures of RRM1 and 2 of CUG-BP1 with 5′-UGUU-3′ are demonstrated (PDB-ID: 3NMR). These figures were generated by the program MOLMOL.21 The protein–RNA complex structures have been elucidated for all of them, except for the HuD C-terminal RRM. Models of the RNA binding modes of (B) HuC/D and (C) CUG-BP1.

Protein–protein interactions in the UHM-type motif. (a) left: SPF45 (ribbon) with the KRKSRWDETP peptide (PDB-ID: 2PEH). Right: SPF45 (surface) with the KRKSRWDETP peptide. (b) The eIF3b RRM (surface) with the DEDVKDNWDDD peptide (PDB-ID: 2KRB). (c) The REF RRM (surface) with the ICP27 peptide (GPLGSVWSRLGARRPSCSP) (PDB-ID: 2KT5). In this figure, only the underlined regions are depicted. In all of the peptide presentations, the side chains of the positively charged, negatively charged, and hydrophobic amino-acid residues are colored blue, red and grey, respectively. These figures were generated by the program MOLMOL.21

Protein–protein interactions on the β-sheet surface. (a) The p14 RRM with the SF3b155 fragment: the main chains of p14 RRM (pink) with the SF3b155 fragment (blue) are demonstrated (PDB-ID: 3LQV). The aromatic side chain of p14 and the SF3b155 peptide are colored grey and purple, respectively. The co-crystallized adenine base is located on RNP2, and is colored pale green. (a) The RRM of the Cyp33 protein with the peptide fragment of the MLL onco-protein (PDB-ID: 2KV7). The main chains of the CYp33 RRM and MLL proteins are colored orange and deep pink, respectively. In addition, their side chains are colored green and magenta, respectively. These figures were generated by the program MOLMOL.21

(a) Dimeric structure of the XePABP2 RRM (PDB-ID: 2JWN). The β-strands of each unit are colored blue and cyan, respectively. Correspondingly, the α-helical regions are colored pink and red. The β2-strands of each unit are paired with each other in an antiparallel manner. (b) The complex structure of FIR RRM1-2 and the N-box peptide of the FBP protein (PDB-ID: 2KXH). The FIR RRM1, RRM2, and the N-box peptide are colored pale green, blue, and deep pink, respectively. These figures were generated by the program MOLMOL.21

Protein–protein interactions involving regions other than the β-sheet surface and the α-helix. (a) The CFIm25/CFIm68 RRM with the target RNA (PDB-ID: 3P6Y). (b) The SF2 RRM2 and SRPK1 (PDB-ID: 3BEJ). (c) The Raver protein with the Vinculin tail domain (PDB-ID: 3H2U). In (a) and (b), hydrophobic interactions primarily mediate the protein–protein interaction. In (c), electrostatic interactions are mainly observed. These figures were generated by the program MOLMOL.21