Eric Condamine

Antimicrobial properties of brevinin-2-related peptide and its analogs: Efficacy against multidrug-resistant Acinetobacter baumannii.

Brevinin‐2 related peptide (B2RP; GIWDTIKSMG10KVFAGKILQN20L.NH2), first isolated from skin secretions of the mink frog Lithobates septentrionalis, shows broad‐spectrum antimicrobial activity but its therapeutic potential is limited by moderate hemolytic activity. The peptide adopts an α‐helical conformation in a membrane‐mimetic solvent but amphipathicity is low. Increasing amphipathicity together with hydrophobicity by the substitutions Lys16→Leu and Lys16→Ala increased hemolytic activity approximately fivefold without increasing antimicrobial potency. The substitution Leu18→Lys increased both cationicity and amphipathicity but produced decreases in both antimicrobial potency and hemolytic activity. In contrast, increasing cationicity of B2RP without changing amphipathicity by the substitution Asp4→Lys resulted in a fourfold increase in potency against Escherichia coli [minimal inhibitory concentration (MIC) = 6 μm) and twofold increases in potency against Staphylococcus aureus (MIC = 12.5 μm) and Candida albicans (MIC = 6 μm) without changing significantly hemolytic activity against human erythrocytes (LC50 = 95 μm). The emergence of antibiotic‐resistant strains of the Gram‐negative bacterium Acinetobacter baumannii constitutes a serious risk to public health. B2RP (MIC = 3–6 μm) and [Lys4]B2RP (MIC = 1.5–3 μm) potently inhibited the growth of nosocomial isolates of multidrug‐resistant Acinetobacter baumannii. Although the analogs [Lys4, Lys18]B2RP and [Lys4, Ala16, Lys18]B2RP showed reduced potency against Staphylococcus aureus, they retained activity against Acinetobacter baumannii (MIC = 3–6 μm) and had very low hemolytic activity (LC50 > 200 μm).

Design of potent, non-toxic antimicrobial agents based upon the naturally occurring frog skin peptides, ascaphin-8 and peptide XT-7.

The frog skin peptides, ascaphin‐8 (GFKDLLKGAAKALVKTVLF.NH2) and XT‐7 (GLLGPLLKIAAKVGSNLL.NH2), show broad‐spectrum antimicrobial activity but their therapeutic potential is limited by toxicity against mammalian cells. Circular dichroism spectra demonstrate that the peptides adopt an amphipathic α‐helical conformation in a membrane‐mimetic solvent. This study has investigated the cytolytic properties of analogs containing selected amino acid substitutions that increase cationicity while maintaining amphipathicity. Substitutions at Ala10, Val14, and Leu18 in ascaphin‐8 by either l‐Lys or d‐Lys produced peptides that retained antimicrobial activity against the bacteria Escherichia coli and Staphylococcus aureus and the opportunistic yeast pathogen, Candida albicans but showed appreciably reduced toxicities (>10‐fold) against human erythrocytes, HepG2 hepatoma‐derived cells, and L929 fibroblasts. The improved therapeutic index of the l‐Lys18 and d‐Lys18 analogs correlated with a decrease in % helicity and in effective hydrophobicity. Substitution of Gly4 by l‐Lys in XT‐7 produced an analog with high potency against micro‐organisms (MIC ≤ 25 μm) but low cytolytic activity against erythrocytes (LD50 > 500 μm) and this increase in therapeutic index also correlated with decreased helicity and hydrophobicity. Analogs of XT‐7 with increased cationicity, containing multiple substitutions by l‐Lys, not only displayed increased antimicrobial potencies, particularly against Candida albicans (MIC ≤ 6 μm), but also increased hemolytic activities.

A glycine-leucine-rich peptide structurally related to the plasticins from skin secretions of the frog Leptodactylus laticeps (Leptodactylidae)

A glycine-leucine-rich peptide was isolated from norepinephrine-stimulated skin secretions of the Sante Fe frog Leptodactylus laticeps (Leptodactylidae) whose primary structure (Gly-Leu-Val-Asn-Gly-Leu-Leu-Ser-Ser-Val-Leu-Gly-Gly-Gly-Gln-Gly-Gly-Gly-Gly-Leu-Leu-Gly-Gly-Ile-Leu) contains the (GXXXG)(3) motif found in the plasticins, previously identified only in phyllomedusid frogs (Hylidae). Circular dichroism studies showed that the secondary structure of the peptide, termed plasticin-L1, was markedly solvent-dependent displaying a random coil conformation in water, a beta-sheet structure in methanol, and an alpha-helical conformation in 50% trifluoroethanol-water. A synthetic replicate of the peptide did not inhibit the growth of Escherichia coli or Staphylococcus aureus or lyse human erythrocytes at concentrations up to 500 microM. At relatively high concentrations (>or=1 microM), the peptide produced a significant (P<0.05), although modest (139% of basal rate at 3 microM), increase in the rate of glucose-induced release of insulin from rat clonal BRIN-BD11 beta cells without increasing the rate of release of lactate dehydrogenase. A peptide, termed ocellatin-L2 was also identified in the skin secretion that was identical to the previously described ocellatin-L1 except for the substitution Asn(23)-->Asp. Ocellatin-L2 was devoid of antimicrobial and hemolytic activity but also showed significant activity in stimulating insulin release from BRIN-BD11 cells (181% of basal rate at 3 microM).

Effect of aminoisobutyric acid (Aib) substitutions on the antimicrobial and cytolytic activities of the frog skin peptide, temporin-1DRa.

Temporin-1DRa (HFLGTLVNLAKKIL.NH(2)), first isolated from the skin of the California red-legged frog Rana draytonii, shows broad-spectrum antimicrobial activity but its therapeutic potential is limited by its toxicity against mammalian cells. The cytolytic properties of cationic alpha-helical peptides are determined by a complex interaction between cationicity, hydrophobicity, conformation, and amphipathicity. This study has investigated the cytolytic properties of conformationally constrained analogs of temporin-1DRa containing alpha-aminoisobutyric acid (Aib) substitutions. Cytolytic activity was determined against the bacteria Escherichia coli and Staphylococcus aureus, the opportunistic yeast pathogen, Candida albicans, human erythrocytes, HepG2 hepatoma-derived cells, and L929 fibroblasts. Aib substitutions at Gly(4), Asn(8), and Ala(10) increased both % helicity, determined in methanol solution, and hydrophobicity resulting in increases in both antimicrobial potencies and toxicities against the mammalian cells. Substitution at Leu(6) resulted in an appreciable decrease in cytolytic activity against all cells whereas the substitutions at His(1), Phe(2), Leu(3), Thr(5), and Val(7) had only minor effects on activity. Substitutions at Leu(9), Ile(13), Leu(14) produced analogs with decreased helicity and hydrophobicity that retained activity against microorganisms but showed appreciably lower cytolytic activities against mammalian cells. In particular, the fourfold increase in therapeutic index [ratio of LC(50) against erythrocytes to minimum inhibitory concentration (MIC) against microorganisms] of [Aib(13)]temporin-1DRa identifies it as a compound with potential for development as a therapeutically valuable anti-infective agent.

Three-dimensional structure of the ion-chanel forming peptide trichorzianin TA VII bound to sodium dodecyl sulfate micelles

Trichorzianin TA VII, Ac0 U1 A2 A3 U4 J5 Q6 U7 U8 U9 S10 L11 U12 P13 V14 U15 I16 Q17 Q18 Fol19, is a nonadecapeptide member of the peptaibol antibiotics biosynthesized by Trichoderma soil fungi, which is characterized by a high proportion of the alpha, alpha-dialkylated amino acids, alpha-aminoisobutyric acid (Aib, U) and isovaline (Iva, J), an acetylated N-terminus and a C-terminal phenylalaninol (Pheol, Fol). The main interest in such peptides stems from their ability to interact with phospholipid bilayers and form voltage-dependent transmembrane channels in planar lipid bilayers. In order to provide insights into the lipid-peptide interaction promoting the voltage gating, the conformational study of TA VII in the presence of perdeuterated sodium dodecyl sulfate (SDS-d25) micelles has been carried out. 1H sequential assignment have been performed with the use of two-dimensional homo- and -heteronuclear nmr techniques including double quantum filtered correlated spectroscopy, homonuclear Hartmann-Hahn, nuclear Overhauser effect spectroscopy, 1H-13C heteronuclear single quantum correlation, and heteronuclear multiple bond correlation. Conformational parameters, such as 3JNHC alpha H coupling constants, temperature coefficients of amide protons (delta gamma/delta TNH) and quantitative nuclear Overhauser enhancement data, lead to detailed structural information. Ninety-eight three-dimensional structures consistent with the nmr data were generated from 231 interproton distances six phi dihedral angle restraints, using restrained molecular dynamics and energy minimization calculations. The average rms deviation between the 98 refined structures and the energy-minimized average structure is 0.59 A for the backbone atoms. The structure of trichorzianin TA VII associated with SDS micelles, as determined by these methods, is characterized by two right-handed helical segments involving residues 1-8 and 11-19, linked by a beta-turn that leads to an angle about 90 degrees-100 degrees between the two helix axes; residues 18 and 19 at the end of the C-terminal helix exhibit multiple conformations.

Synthesis and Structure of New 3,3,9,9-Tetrasubstituted- 2,4,8,10-Tetraoxaspiro(5.5)undecane Derivatives

The configurational and conformational behavior of some new 3,3,9,9-tetra-substituted-2,4,8,10-tetraoxaspiro[5.5]undecane derivatives with axial chirality was investigated by conformational analysis and variable temperature NMR experiments.

Ring-chain tautomerism and other versatile behaviour of 1,4-diimino- and 1,2-phenylene derivatives of some C-substituted serinols

Abstract The behaviour of the title compounds, obtained from terephthaldicarboxaldehyde or 1,2-phthaldicarboxaldehyde and certain (non)chiral C -substituted serinols is discussed in terms of ring (chain)–chain (ring) tautomerism (1,4-phenylene derivatives) or formal intramolecular redox isomerisation (1,2-derivatives).

New [4.4]cyclophanes: molecular parallelograms, triangles, rhombuses, pentagons, and supramolecular constructions.

The fair or good yield synthesis of new [(4.4)(n)]cyclophanes (n = 1-5), starting from 1,4-bis(2-hydroxymethyl-5,5-dimethyl-1,3-dioxan-2-yl)benzene and several diacid-dichlorides, based on monomer and oligomer formation reactions (from 1 + 1 to 5 + 5), is reported. The structure and the complex architectures of the lattices for these cyclophanes are revealed by the X-ray molecular structure for five compounds, NMR investigations, and mass spectrometry measurements. Intramolecular and intermolecular CH-pi, p-pi, and pi-pi interactions are observed, both in solid state and solution.

Structural and pharmacological characteristics of chimeric peptides derived from peptide E and β-endorphin reveal the crucial role of the C-terminal YGGFL and YKKGE motifs in their analgesic properties

Peptide E (a 25-amino acid peptide derived from proenkephalin A) and beta-endorphin (a 31-amino acid peptide derived from proopiomelanocortin) bind with high affinity to opioid receptors and share structural similarities but induce analgesic effects of very different intensity. Indeed, whereas they possess the same N-terminus Met-enkephalin message sequence linked to a helix by a flexible spacer and a C-terminal part in random coil conformation, in contrast with peptide E, beta-endorphin produces a profound analgesia. To determine the key structural elements explaining this very divergent opioid activity, we have compared the structural and pharmacological characteristics of several chimeric peptides derived from peptide E and beta-endorphin. Structures were obtained under the same experimental conditions using circular dichroism, computational estimation of helical content and/or nuclear magnetic resonance spectroscopy (NMR) and NMR-restrained molecular modeling. The hot-plate and writhing tests were used in mice to evaluate the antinociceptive effects of the peptides. Our results indicate that neither the length nor the physicochemical profile of the spacer plays a fundamental role in analgesia. On the other hand, while the functional importance of the helix cannot be excluded, the last 5 residues in the C-terminal part seem to be crucial for the expression or absence of the analgesic activity of these peptides. These data raise the question of the true function of peptides E in opioidergic systems.

Design, Synthesis and Structural Analysis of New Macrocycles Containing Dispiro-1,3-dioxane Units

The synthesis and the structure of new macrocycles containing semiflexible dispiro-1,3-dioxane units is reported. The structural analysis of the compounds is performed by high field NMR spectra, mass spectrometry investigations (MALDI, ESI-MS) and the solid state molecular structure obtained for two compounds by single crystal X-ray diffractometry. The dynamics of the macrocycles promoted by the flipping of the middle cyclohexane ring of the dispirane units is investigated using low temperature NMR experiments. New macrocycles containing dispiro-1,3-dioxane units were investigated by NMR, X-ray diffractometry and mass spectrometry