Toward the Molecular Basis of Inherited Prion Diseases: NMR Structure of the Human Prion Protein with V210I Mutation Ivana Biljan 1 †‡, Gregor Ilc 1, 2 †, Gabriele Giachin 3 †, Andrea Raspadori 3 , Igor Zhukov 1, 2 §, Janez Plavec 1, 2, 4 ⁎ and Giuseppe Legname 3, 5, 6 ⁎ 1 Slovenian NMR Centre, National Institute of Chemistry, Hajdrihova 19, SI-1000 Ljubljana, Slovenia 2 EN-FIST Centre of Excellence, Dunajska 156, SI-1001 Ljubljana, Slovenia 3 Laboratory of Prion Biology, Neurobiology Sector, Scuola Internazionale Superiore di Studi Avanzati (SISSA), Via Bonomea 265, I-34136 Trieste, Italy 4 Faculty of Chemistry and Chemical Technology, University of Ljubljana, Aškerčeva cesta 5, SI-1000 Ljubljana, Slovenia 5 Italian Institute of Technology, SISSA Unit, Via Bonomea 265, I-34136 Trieste, Italy 6 ELETTRA Laboratory, Sincrotrone Trieste S.C.p.A., Basovizza, I-34149 Trieste, Italy Received 25 June 2011; received in revised form 28 July 2011; accepted 28 July 2011 Available online 4 August 2011 Edited by P. Wright Keywords: prions; mutants; transmissible spongiform encephalopathies; genetic Creutzfeldt–Jakob disease; NMR structure determination The development of transmissible spongiform encephalopathies (TSEs) is associated with the conversion of the cellular prion protein (PrP C ) into a misfolded, pathogenic isoform (PrP Sc ). Spontaneous generation of PrP Sc in inherited forms of disease is caused by mutations in gene coding for PrP (PRNP). In this work, we describe the NMR solution-state structure of the truncated recombinant human PrP (HuPrP) carrying the pathological V210I mutation linked to genetic Creutzfeldt–Jakob disease. The three-dimen- sional structure of V210I mutant consists of an unstructured N-terminal part (residues 90–124) and a well-defined C-terminal domain (residues 125–228). The C-terminal domain contains three α-helices (residues 144–156, 170–194 and 200–228) and a short antiparallel β-sheet (residues 129–130 and 162– 163). Comparison with the structure of the wild-type HuPrP revealed that although two structures share similar global architecture, mutation introduces some local structural differences. The observed variations are mostly clustered in the α 2 –α 3 inter-helical interface and in the β 2 –α 2 loop region. Introduction of bulkier Ile at position 210 induces reorientations of several residues that are part of hydrophobic core, thus influencing α 2 –α 3 *Corresponding authors. J. Plavec is to be contacted at Slovenian NMR Centre, National Institute of Chemistry, Hajdrihova 19, SI-1000 Ljubljana, Slovenia; G. Legname, Laboratory of Prion Biology, Neurobiology Sector, Scuola Internazionale Superiore di Studi Avanzati (SISSA), Via Bonomea 265, I-34136 Trieste, Italy. E-mail addresses: janez.plavec@ki.si; giuseppe.legname@sissa.it. † I.B., G.I. and G.G. contributed equally to the work. ‡ I.B. is on leave from Department of Chemistry, Faculty of Science, University of Zagreb, Horvatovac 102A, HR-10000 Zagreb, Croatia. § I.Z. is on leave from Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Pawinskiego 5A, 02-106 Warsaw, Poland. Abbreviations used: WT, wild type; HSQC, heteronuclear single quantum coherence; NOE, nuclear Overhauser enhancement; NOESY, NOE spectroscopy; TOCSY, total correlated spectroscopy; 3D, three-dimensional; TSE, transmissible spongiform encephalopathy; HuPrP, human PrP; CJD, Creutzfeldt–Jakob disease; GSS, Gerstmann–Sträussler–Scheinker; fCJD, familial CJD; TEV, tobacco etch virus; EDTA, ethylenediaminetetraacetic acid; PDB, Protein Data Bank. doi:10.1016/j.jmb.2011.07.067 J. Mol. Biol. (2011) 412, 660–673 Contents lists available at www.sciencedirect.com Journal of Molecular Biology journal homepage: http://ees.elsevier.com.jmb 0022-2836/$ - see front matter © 2011 Elsevier Ltd. All rights reserved.