Ettore Benedetti

Intramolecularly Hydrogen-Bonded Peptide Conformation

Over the past few years the possible occurrence of intramolecularly hydrogen-bonded structures in linear and cyclic peptides has attracted increasing attention. In this review emphasis is given to solid-state studies, particularly by X-ray diffraction and infrared absorption techniques. Conformational energy calculations are also considered. The discussion is focused both on model peptides and biological activity polypeptide molecules. The tetrapeptide system (Formula: see text), examined allows one to discuss the extended C5 structure and the various folded conformations, namely the C7 (gamma-turn), C8, C10 (beta-turn), C11, and C13 conformations. The four latter forms may include cis peptide configurations. The oxy-analogs to the C7, C10, and C13 conformations and structures containing bifurcated hydrogen bonds are also discussed. The last sections describe intramolecularly hydrogen-bonded peptide structures involving: (1) a side-chain group, (2) the N-protecting group (in synthetic model compounds), and (3) a beta-amino acid.

Peptide-based Molecules in Angiogenesis

Angiogenesis refers to the process of remodeling the vascular tissue characterized by the branching out of a new blood vessel from a pre‐existing vessel. Angiogenesis is particularly active during embryogenesis, while during adult life it is quiescent and limited to particular physiologic phenomena. Recently, the study of molecular mechanisms of angiogenesis has stirred renewed interest due to the recognition of the role played by angiogenesis in several pathologies of significant medical impact, such as cancer and cardiovascular disease, and due to the pharmacologic interest rising from the possibility of modulating these phenomena. Antibodies, peptides and small molecules targeting active endothelial cells represent an innovative tool in therapeutic and diagnostic fields. In this study, we reviewed the literature of peptide and peptidomimetics in angiogenesis and their potential applications. Two specific protein systems, namely the vascular endothelial growth factor and its receptor and integrins, will be discussed in detail.

Critical main-chain length for conformational conversion from 3(10)-helix to alpha-helix in polypeptides.

To assess the minimal peptide length required for the stabilization of the alpha-helix relative to the 3(10)-helix in Aib-rich peptides, we have solved the X-ray diffraction structures of the terminally blocked sequential hexa- and octapeptides with the general formula-(Aib-L-Ala)n-(n = 3 and 4, respectively). The hexapeptide molecules are completely 3(10)-helical with four 1----4 intramolecular N-H . . . O = C H-bonds. On the other hand, the octapeptide molecules are essentially alpha-helical with four 1----5 H-bonds; however, the helix is elongated at the N-terminus, with two 1----4 H-bonds, giving these molecules a mixed alpha/3(10)-helical character. In both compounds the right-handed screw sense of the helix is dictated by the presence of the Ala residues of L-configuration. This study represents the first experimental proof for a 3(10)----alpha-helix conversion in the crystal state induced by peptide backbone lengthening only.

The longest, regular polypeptide 310 helix at atomic resolution

A synthetic, terminally blocked homodecapeptide from the C alpha, alpha-dimethylated glycyl residue alpha-aminoisobutyric acid has been analyzed by single-crystal X-ray diffraction and the structure refined to R = 0.073. The compound crystallizes as a perfect 3(10) helix, stabilized by eight consecutive intramolecular N-H . . . O = C hydrogen bonds. This is the first observation at atomic resolution of a regular polypeptide 3(10) helix as long as three complete turns.

Nitroxyl peptides as catalysts of enantioselective oxidations.

The achiral, nitroxyl-containing alpha-amino acid TOAC (TOAC = 2,2,6,6-tetramethylpiperidine-1-oxyl-4-amino-4-carboxylic acid), in combination with the chiral alpha-amino acid C(alpha)-methyl valine [(alphaMe)Val], was used to prepare short peptides (from di- to hexa-) that induced the enantioselective oxidation of racemic 1-phenylethanol to acetophenone. The best catalyst was an N(alpha)-acylated dipeptide alkylamide with the -TOAC-(alphaMe)Val- sequence folded in a stable, intramolecularly hydrogen-bonded beta-turn conformation with large, lipophilic (hydrophobic) N- and C-terminal blocking groups. We rationalized our findings by proposing models for the diastereomeric intermediates between (R)-[and (S)]-1-phenylethanol and the catalyst Fmoc-TOAC-L-(alphaMe)Val-NHiPr, based on the X-ray diffraction structure of the latter.

Gadd45β forms a Homodimeric Complex that Binds Tightly to MKK7

Gadd45 alpha, beta, and gamma proteins, also known as growth arrest and DNA damage-inducible factors, have a number of cellular functions, including cell-cycle regulation and propagation of signals produced by a variety of cellular stimuli, maintaining genomic stability and apoptosis. Furthermore, Gadd45 beta has been indicated as a major player in the endogenous NF-kappaB-mediated resistance to apoptosis in a variety of cell lines. In fibroblasts this mechanism involves the inactivation of MKK7, the upstream activator of JNK, by direct binding within the kinase ATP pocket. On the basis of a number of experimental data, the structures of Gadd45 beta and the Gadd45 beta-MKK7 complex have been predicted recently and data show that interactions are mediated by acidic loops 1 and 2, and helices 3 and 4 of Gadd45 beta. Here, we provide further evidence that Gadd45 beta is a prevailingly alpha-helical protein and that in solution it is able to form non covalent dimers but not higher-order oligomers, in contrast to what has been reported for the homologous Gadd45 alpha. We show that the contact region between the two monomers is comprised of the predicted helix 1 (residues Q17-Q33) and helix 5 (residues K131-R146) of the protein, which appear to be antiparallel and to form a large dimerisation surface not involved in MKK7 recognition. The results suggest the occurrence of a large complex containing at least an MKK7-Gadd45 beta:Gadd45 beta-MKK7 tetrameric unit whose complexity could be further increased by the dimeric nature of the isolated MKK7.

Preferred conformation of peptides rich in alicyclic Cα,α-disubstituted glycines

The conformational preferences of the alicyclic Cα,α‐disubstituted glycines Acnc (1‐amino‐1‐cycloalkane‐carboxylic acid; n = 4, 7, 9, 12) were assessed in selected model compounds, including homopeptides and Ala (or Aib, α‐aminoisobutyric acid)/Acnc peptides containing a small total number of residues, by Fourier transform ir absorption, 1H‐nmr, and x‐ray diffraction analyses. The results obtained indicate that β‐turn and 310‐helical structures are preferentially adopted by short peptides rich in these cycloaliphatic α‐amino acids.

Micelles derivatized with octreotide as potential target-selective contrast agents in MRI†‡

New amphiphilic monomers (OCA‐DTPAGlu and OCA‐DOTA) containing, in the same molecule, three different functions: (i) the chelating agent (DTPAGlu or DOTA) able to coordinate gadolinium ion, (ii) the octreotide bioactive peptide able to target somatostatin receptors, and (iii) a hydrophobic moiety with two 18‐carbon atoms alkyl chains have been designed and synthesized by solid‐phase methods. The novel amphiphilic monomers aggregate, in water solution, giving stable micelles at very low concentration (cmc values of 2.3 × 10−6 mol kg−1 and 2.5 × 10−6 mol kg−1 for OCA‐DTPAGlu and OCA‐DOTA, respectively) as confirmed by fluorescence spectroscopy. Fluorescence studies and circular dichroism experiments indicate, for the two compounds as well as for their gadolinium complexes (OCA‐DOTA(Gd) and OCA‐DTPAGlu(Gd)), the complete exposure of octreotide on the micelle surface, and the predominant presence of an antiparallel β‐sheet peptide conformation characterized by a β‐like turn. The high relaxivity value (r1p = 13.9 mM−1 s−1 at 20 MHz and 25 °C), measured for micelles obtained by the gadolinium complex OCA‐DTPAGlu(Gd), indicates these aggregates as promising target‐selective magnetic resonance imaging (MRI) contrast agents. Copyright

Peptides containing membrane-interacting motifs inhibit herpes simplex virus type 1 infectivity

Abstract Herpes simplex virus (HSV) membrane fusion represents an attractive target for anti-HSV therapy. To investigate the structural basis of HSV membrane fusion and identify new targets for inhibition, we have investigated the different membranotropic domains of HSV-1 gH envelope glycoprotein. We observed that fusion peptides when added exogenously are able to inhibit viral fusion likely by intercalating with viral fusion peptides upon adopting functional structure in membranes. Interestingly, peptides analogous to the predicted HSV-1 gH loop region inhibited viral plaque formation more significantly. Their inhibitory effect appears to be a consequence of their ability to partition into membranes and aggregate within them. Circular dichroism spectra showed that peptides self-associate in aqueous and lipidic solutions, therefore the inhibition of viral entry may occur via peptides association with their counterpart on wild-type gH. The antiviral activity of HSV-1 peptides tested provides an attractive basis for the development of new fusion peptide inhibitors corresponding to regions outside the fusion protein heptad repeat regions.

Regularly alternatingL,D-peptides. II. The double-stranded right-handed antiparallel ?-helix in the structure oft-Boc-(L-Phe-D-Phe)4-OMe

The crystal structure of Boc‐(L‐Phe‐D‐Phe)4‐OMe has been determined by x‐ray diffraction analysis. The peptide crystallizes in the triclinic system, space group P1 with a = 15.290 Å, b = 15.163 Å, c = 19.789 Å, α = 102.49°, β = 96.59°, γ = 74.22°, and Z = 2. The structure has been solved by coupling of the molecular replacement technique and expansion by tangent formula refinement of the set of known phases. Several cycles of Fourier calculations and least‐squares refinement led to the location of 194 atoms of the two independent octapeptide chains and few molecules of cocrystallized solvent (chloroform, water, and methanol). The isotropic refinement converged to R = 0.13 for the 3077 “observed” reflections.