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Proceedings of the 31 st European Peptide SymposiumMichal Lebl, Morten Meldal, Knud J. Jensen, Thomas Hoeg-Jensen (Editors)European Peptide Society, 2010Synthesis, Preferred Conformation, and Membrane Activity ofHeptaibin, a Medium-Length PeptaibioticMarta De Zotti, Barbara Biondi, Cristina Peggion, Marco Crisma,Fernando Formaggio, and Claudio TonioloICB, Padova Unit, CNR, Department of Chemistry, University of Padova,Padova, 35131, ItalyIntroductionThe medium-length peptaibiotics [1] are characterized by a primary structure of 13-15amino acid residues and include inter alia some samarosporins, stilbellins, bergofungins,and emerimicins. Despite the interesting antibiotic and antifungal properties exhibited bythese membrane-active peptides, their exact mechanism of action is still unknown. Ourcurrent aim is to investigate the relationships between their conformational properties andbioactivity.Results and DiscussionHere, we present our results on heptaibin, a 14-amino acid residue peptaibiotic, extractedfrom the culture of Emericellopsis sp. BAUA8289 and chemically characterized byIshiyama et al. ten years ago [2]. The heptaibin primary structure is as follows:Ac-Phe-(Aib) 3 -Val-Gly-Leu-(Aib) 2 -Hyp-Gln-Aib-Hyp-Aib-Phol(Ac, acetyl; Aib, -aminoisobutyric acid; Hyp, 4(R)-hydroxy-(S)-proline; Phol, the1,2-aminoalcohol (S)-phenylalaninol). The solid-phase synthesis of bergofungin D, thesequence of which is similar to that of heptaibin, has been recently reported [3].Our solid-phase synthesis of heptaibin involved the Fmoc-protection/HATUC-activation methodology. Special attention was devoted to prevent 2,5-dioxopiperazineformation, particularly when the N-terminal sequence of the growing chain is the H-Aib-Hyp- dipeptide. The final product was purified (Figure 1) and fully characterized. Adetailed conformational analysis was performed by use of FT-IR absorption, CD, 2D-NMRcombined with molecular dynamics (MD) calculations, and X-ray crystallography (thelatter technique on an N-terminal segment). Being rich (>50%) of the non-codedC -tetrasubstituted Aib residue, heptaibin is mostly helical under different experimentalconditions (Figure 2) and it is stable against the pronase E proteolytic attack. Fluorescenceleakage experiments revealed that heptaibin is a membrane-permeabilizing compound.15.8 min0 10 20 30time (min)Fig. 1. RP-HPLC profile obtained for heptaibin. Column: Vydac C 18 ; gradient: 50-100%B in 25min; eluants: A: H 2 O+0.1% TFA, B: CH 3 CN/H 2 O 9:1+0.1% TFA.392