Elena Nardi

A synthetic glycopeptide of human myelin oligodendrocyte glycoprotein to detect antibody responses in multiple sclerosis and other neurological diseases

Glycopeptides of hMOG(30-50) containing a glucosyl moiety on the side-chains of Asn, Ser or Hyp at position 31 were synthesised. Antibody titres to hMOG(30-50) and to its glucoderivatives were measured by ELISA in sera of patients affected by different neurological diseases. Anti-hMOG(30-50) antibodies were detected only using the glycopeptide [Asn31(N-Glc)]hMOG(30-50).

Diastereoselective Alkylation of Schiff Bases for the Synthesis of Lipidic Unnatural Fmoc-Protected α-Amino Acids

Peptides with increased lipophilicity can cross cell membranes more easily and have longer half-life times. For these reasons, the synthesis of enantiomerically pure Fmoc-protected lipidic α-amino acids is a relevant goal. Schiff bases originating from the reaction between the two enantiomers of 2-hydroxypinan-3-one with Gly-OtBu were alkylated with a series of long alkyl halides. Diastereomeric excesses were determined by reversed-phase HPLC, under conditions carefully chosen for such lipophilic substrates. (© Wiley-VCH Verlag GmbH, 69451 Weinheim, Germany, 2002)

Structural characterization of lipopeptide agonists for the bradykinin B2 receptor

The conformational features of Pam-Lys(0)-Arg(1)-Pro(2)-Pro(3)-Gly(4)-Phe(5)-Ser(6)-Pro(7)-Phe(8)-Arg(9)-OH (PKD) and Pam-Gly(-1)-Lys(0)-Arg(1)-Pro(2)-Pro(3)-Gly(4)-Phe(5)-Ser(6)-Pro(7)-Phe(8)-Arg(9)-OH (PGKD), the Pam-Lys and Pam-Gly-Lys analogues of bradykinin, have been determined by high-resolution NMR in a zwitterionic lipoid environment. Radical-induced relaxation of the (1)H NMR signals was used to probe the topological orientation of the peptides with respect to the zwitterionic lipid interface. The radical-induced relaxation and molecular dynamics (MD) data indicated that the palmitic acid and N-terminal amino acid residues embed into the micelles, while the rest of the polypeptide chain is closely associated with the water-micelle interface. Throughout the entire nuclear Overhauser effect restrained MD simulation, a nonideal type I beta-turn was observed in the C-terminus of PKD between residues 6 and 9, and a gamma-turn was observed in the C-terminus of PGKD between residues 6 and 7. Therefore, the additional glycine has a dramatic effect on the structural preferences of the biologically important C-terminus, an effect brought about by the interaction with the lipid environment. These structural features are correlated to the biological activity at the bradykinin B2 receptor.

Synthesis of lipopeptides of the immunodominant epitope hMBP(83–99) containing amide or C-C bond linked hydrophobic chains for the study of T cell response

We previously demonstrated that the lipopeptide of the myelin basic protein (MBP) immunodominant epitope in Lewis rat Palm-GpMBP(74-85) (Gp: guinea pig), which induced experimental autoimmune encephalomyelitisin vivo strongly increased the T cell proliferative responsein vitro. We extended this study to the human immunodominant epitope hMBP(83-99), synthesizing different lipophilic peptides bearing a hydrophobic chain linked through an amide or a C-C bond. To this aim, we developed a synthetic pathway for (±)-N-Fmoc-Ahd-OH (Ahd: 2-aminohexadecanoic acid) which was used to synthesize diastereomeric peptides which were successfully separated by reverse-phase high-performance liquid chromatography. MBP-specific T cell lines recognizing the immunodominant epitope hMBP(83-99) have been generated from patients affected by multiple sclerosis. Their proliferative response to the native peptide and to some lipoderivatives has been investigated. In contrast to the animal model, none of the investigated lipopeptides exhibited ‘superagonist’ activity.

Recombinant MOG from baculovirus inhibits anti-hMOG(30-50) antibodies detected by the synthetic antigen [Asn31(Glc)]hMOG(30-50)

Elena Nardi, Silvia Mazzucco, Sabrina Mata, Mario Chelli, Benedetta Mazzanti, Elisabetta Traggiai, Mauro Ginanneschi, Francesco Pinto, Luca Massacesi, Marco Vergelli, Hubert Kalbacher, Francesco Lolli, and Anna M. Papini Dipartimento di Chimica Organica “Ugo Schiff” and C.N.R. CSCEA, Universita degli Studi di Firenze, I-50121 Firenze, Italy; Servizio di Neurofisiopatologia, Dipartimento di Scienze Neurologiche e Psichiatriche, and Dipartimento di Scienze Neurologiche e Psichiatriche, Universita degli Studi di Firenze, I-50134 Firenze, Italy; and Medizinisch und Naturwissenschaftliches Forschungszentrum, Universitat Tubingen, D-72074 Tubingen, Germany.

Conformational Studies of a Glycopeptide Recognized with High Affinity by Autoantibodies in Multiple Sclerosis

Myelin oligodendrocyte glycoprotein (MOG), a minor myelin component, is an important central-nervous system-specific target autoantigen for primary demyelination in autoimmune diseases like multiple sclerosis (MS). It has been described that glycosylation of a MOG peptide epitope improved the detection of specific autoantibodies in sera of MS patients [1]. We have recently found that the specific antibody recognition of this peptide is most likely driven by direct interactions of the antibody binding site with the Asn-linked sugar moiety and then stabilized by putative specific peptide-antibody interactions [2]. We have subsequently prepared and characterized a new 21-mer MOG derived glycopeptide N7 (H-Thr-Pro-Arg-Val-Glu-Arg-Asn(Glc)-Gly-His-Ser-Val-Phe-Leu-Ala-Pro-Tyr-Gly-Trp-Met-Val-Lys-OH — 1) showing enhanced affinity for MS autoantibodies in ELISA experiments compared to the native N-glycosylated MOG peptide epitope Asn31(Glc)hMOG(30–50). Thus, the peptide (1) is a synthetic antigen able to detect pathogenic demyelinating autoantibodies in MS patients and, most importantly, it can be used as a template for the design of a new generation of drugs capable of specific blockage of circulating autoantibodies in patients affected by MS. In view of this goal, we describe here a conformational analysis of this peptide, and of its unglycosylated and inactive counterpart (H-Thr-Pro-Arg-Val-Glu-Arg-Asn-Gly-His-Ser-Val-Phe-Leu-Ala-Pro-Tyr-Gly-Trp-Met-Val-Lys-OH — 2).