1 3 DOI 10.1007/s00726-014-1841-9 Amino Acids (2014) 46:2841–2853 ORIGINAL ARTICLE Structure of the cyclic peptide [W8S]contryphan Vn: effect of the tryptophan/serine substitution on trans–cis proline isomerization Ridvan Nepravishta · Walter Mandaliti · Sonia Melino · Tommaso Eliseo · Maurizio Paci Received: 25 April 2014 / Accepted: 17 September 2014 / Published online: 27 September 2014 © Springer-Verlag Wien 2014 Keywords Contryphan · NMR · Cyclic peptides · Three-dimensional structure · Fluorescence · Proline isomerization Abbreviations TFA Trifluoroacetic acid HMQC Heteronuclear multiple quantum coherence HSQC Heteronuclear single quantum coherence ROESY Rotating-frame Overhauser spectroscopy TOCSY Total correlation spectroscopy DOSY Diffusion-ordered spectroscopy TFE Trifluoroethanol PME Particle-mesh Ewald method TSP Trimethylsilyl propionic acid Introduction Contryphans are natural bioactive peptides, first isolated from piscivorous cone snail venom (Olivera et al. 1985; Jimenez et al. 1996; Jacobsen et al. 1998); all of them have S–S cystine bridges and are rich in a variety of unusual post-translational modifications, including proline hydrox- ylation, C-terminus amidation, proline and leucine isomeri- zation (Jacobsen et al. 1999; Pallaghy et al. 1999) and tryptophan bromination (Jimenez et al. 1997). The charac- terized Contryphans are reported schematically in Table 1 (Olivera et al. 1985; Jimenez et al. 1996, 1997, 2001, 2002; Jacobsen et al. 1998, 1999; Pallaghy et al. 1999, 2000; Pal- laghy and Norton 2000; Massilia et al. 2001, 2003; Eliseo et al. 2004; Grant et al. 2004; Hansson et al. 2004; Saba- reesh et al. 2006). The biological activity of Contryphans has been assayed (Jimenez et al. 1996, 2001; Massilia et al. 2001); specifically in the case Contryphan Vn, it was found to modulate the activity of Ca 2+ -dependent K + channels Abstract The structural characterization of [W8S] contryphan Vn, an analogue of Contryphan Vn with tryp- tophan 8 substituted with a serine residue (W8S), was performed by NMR spectroscopy, molecular dynamics simulations and fluorescence spectroscopy. Contryphan Vn, a bioactive cyclic peptide from the venom of the cone snail Conus ventricosus, contains an S–S bridge between two cysteines and a D-tryptophan. Like other Contryphans, [W8S]contryphan Vn has proline 7 isomerized trans, while the proline 4 has nearly equivalent populations of cis and trans configurations. The thermodynamic and kinetic parameters of the trans–cis isomerization of proline 4 were measured. The isomers of [W8S]contryphan Vn with proline 4 in cis and trans show structural differences. The absence of the salt bridge between the same Asp2 and Lys6, present in Contryphan Vn, may be attributed to the lack of the hydrophobic side chain of Trp8 where it likely protects the electrostatic interactions. These results may contribute to identifying, in these cyclic peptides, the structural deter- minants of the mechanism of proline trans–cis isomeriza- tion, this being also an important step in protein folding. R. Nepravishta and W. Mandaliti have equally contributed to this work. Electronic supplementary material The online version of this article (doi:10.1007/s00726-014-1841-9) contains supplementary material, which is available to authorized users. R. Nepravishta · W. Mandaliti · S. Melino · T. Eliseo · M. Paci (*) Department of Chemical Sciences and Technologies, University of Rome “Tor Vergata”, Via della Ricerca Scientifica 1, 00133 Rome, Italy e-mail: paci@uniroma2.it