Homer EVH1 (Ena/VASP Homology 1) domains interact with proline-rich motifs in the cytoplasmic regions of group 1 metabotropic glutamate receptors (mGluRs), inositol-1,4,5-trisphosphate receptors (IP3Rs), and Shank proteins. We have determined the crystal structure of the Homer EVH1 domain complexed with a peptide from mGluR (TPPSPF). In contrast to other EVH1 domains, the bound mGluR ligand assumes an unusual conformation in which the side chains of the Ser-Pro tandem are oriented away from the Homer surface, and the Phe forms a unique contact. This unusual binding mode rationalizes conserved features of both Homer and Homer ligands that are not shared by other EVH1 domains. Site-directed mutagenesis confirms the importance of specific Homer residues for ligand binding. These results establish a molecular basis for understanding the biological properties of Homer-ligand complexes.

Bulged-G motifs are ubiquitous internal RNA loops that provide specific recognition sites for proteins and RNAs. To establish the common and distinctive features of the motif we determined the structures of three variants and compared them with related structures. The variants are 27-nt mimics of the sarcin/ricin loop (SRL) from Escherichia coli 23S ribosomal RNA that is an essential part of the binding site for elongation factors (EFs). The wild-type SRL has now been determined at 1.04 A resolution, supplementing data obtained before at 1.11 A and allowing the first calculation of coordinate error for an RNA motif. The other two structures, having a viable (C2658U*G2663A) or a lethal mutation (C2658G*G2663C), were determined at 1.75 and 2.25 A resolution, respectively. Comparisons reveal that bulged-G motifs have a common hydration and geometry, with flexible junctions at flanking structural elements. Six conserved nucleotides preserve the fold of the motif; the remaining seven to nine vary in sequence and alter contacts in both grooves. Differences between accessible functional groups of the lethal mutation and those of the viable mutation and wild-type SRL may account for the impaired elongation factor binding to ribosomes with the C2658G*G2663C mutation and may underlie the lethal phenotype.

NM23-H2, a presumed regulator of tumor metastasis in humans, is a hexameric protein with both enzymatic (NDP kinase) hexamers, suggesting that they perform at the level of DNA recognition and that separate functional domains exist for enzyme catalysis and DNA binding. In the context of the known crystal structure of NM23-H2, the DNA-binding residues are located within distinct structural motifs in the monomer, which are exposed to the surface near the 2-fold axis of adjacent subunits in the hexamer. These findings are explained by a model in which NM23-H2 binds DNA with a combinatorial surface consisting ofthe "outer" face ofthe dimer. Chemical crosslinking data support a dimeric DNA-binding mode by

Sarcin is a member of a fungal toxin family that enters cells and specifically cleaves one of the thousands of RNA phosphodiester bonds in the ribosome. As a result, elongation factor binding is disrupted, translation is inhibited and apoptosis is triggered. The toxin targets a universal RNA structure in the ribosome called the sarcin/ricin loop (SRL). A 1.11 Å resolution structure of a minimal SRL RNA substrate (∼30‐mer) shows that the loop portion of the substrate folds into two common building blocks of RNA structure: a bulged‐G motif (recognition site) and a GAGA tetraloop (cleavage site). To elucidate the structural basis of toxin action, two co‐crystal structures of the sarcin homologue restrictocin have beeen determined bound to different analogs of a minimal SRL RNA substrate. Our studies argue that site selection by the toxin depends on direct base and shape recognition of the SRL RNA, and that cleavage by the toxin depends on a base flipping mechanism that positions the nucleophile for in‐line attack on the scissile bond.

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