C OMMUNICATION Solution Structure of the Vts1 SAM Domain in the Presence of RNA Thomas A. Edwards 1 ‡, Joel A. Butterwick 2 ‡, Lei Zeng 1 , Yogesh K. Gupta 1 Xin Wang 1 , Robin P. Wharton 3 , Arthur G. Palmer III 2 * and Aneel K. Aggarwal 1 * 1 Structural Biology Program Department of Physiology and Biophysics, Mount Sinai School of Medicine, Box 1677, 1425 Madison Avenue, New York NY 10029, USA 2 Department of Biochemistry and Molecular Biophysics Columbia University, 630 West 168th Street, Box 36, New York NY 10032, USA 3 Howard Hughes Medical Institute, Department of Molecular Genetics and Microbiology, Box 3657, Duke University Medical Center Durham, NC 27710, USA The yeast Vts1 SAM (sterile alpha motif) domain is a member of a new class of SAM domains that specifically bind RNA. To elucidate the structural basis for RNA binding, the solution structure of the Vts1 SAM domain, in the presence of a specific target RNA, has been solved by multidimensional heteronuclear NMR spectroscopy. The Vts1 SAM domain retains the “core” five-helix-bundle architecture of traditional SAM domains, but has additional short helices at N and C termini, comprising a small substructure that caps the core helices. The RNA- binding surface of Vts1, determined by chemical shift perturbation, maps near the ends of three of the core helices, in agreement with mutational data and the electrostatic properties of the molecule. These results provide a structural basis for the versatility of the SAM domain in protein and RNA-recognition. q 2005 Elsevier Ltd. All rights reserved. Keywords: Vts1; SAM domain; Smaug; RNA-binding protein; translational control *Corresponding authors The SAM (sterile alpha motif) domain is a common protein–protein interaction module, identified in a wide variety of eukaryotic proteins, including the signal transducing EphB2 and EphA4 receptors, the transcriptional repres- sors TEL and polyhomeotic (Ph), and the splice variants (p73 and p63) of the tumor suppressor p53. 1–6 Recently, we and others showed that the SAM domain of Smaug (Smg), a protein required for proper abdominal segmentation in early Drosophila melanogaster embryos, encodes a novel protein–RNA, rather than a traditional protein– protein, interaction module. 7,8 Smg acts during early Drosophila development to repress the translation of maternal nanos (nos) mRNA by binding to a 23 nt RNA hairpin, consisting of a nine base-pair stem and a CUGGC loop, within the so-called TCE (trans- lation control element) of the 3 0 untranslated region of nos mRNA. 9,10 The mechanism by which Smg represses translation is not yet defined, although it may involve interference with both pre- and post-initiation events. 11,12 In addition, Smg has been shown to play a role in the degradation of maternal Hsp83 mRNA. 13 Although structures are known for several SAM domains that mediate protein–protein inter- actions (as homo- and hetero-typic oligomers), 6 the only structure known for a SAM domain that mediates protein–RNA interactions is the Smg 0022-2836/$ - see front matter q 2005 Elsevier Ltd. All rights reserved. Present address: T. A. Edwards, Astbury Centre for Structural Molecular Biology, Institute of Molecular and Cellular Biology, University of Leeds, Leeds LS2 9JT, UK. ‡ T.A.E. & J.A.B. contributed equally to this work. Abbreviations used: SAM, sterile alpha motif; TCE, translation control element; NOE, nuclear Overhauser effect; HSQC, heteronuclear single quantum coherence; PHAT, pseudo-HEATanalogous topology; RBD, RNA-binding domain; Smg, Smaug. E-mail addresses of the corresponding authors: agp6@columbia.edu; aggarwal@inka.mssm.edu doi:10.1016/j.jmb.2005.12.004 J. Mol. Biol. (2006) 356, 1065–1072