The crystal and molecular structure of cyclo-[-(D-Val-Hyi-Val-D-Hyi)3-] (meso-valinomycin,C60H102N6O18)

The crystal structure of the valinomycin analog, cyclo-[(-D -Val-Hyi-Val-D -Hyi-)₃-] (meso-valinomycin, C₆₀H₁₀₂N₆O₁₈) has been determined by direct x-ray diffraction procedures. The crystals are triclinic, space group P1 , number of molecules per unit cell Z = 1, and cell parameters a = 11.831, b = 13.815, c = 14.889 Å, α = 109.54°, β = 116.10°, γ = 98.89°. The atomic coordinates for the C,N,O atoms were refined in the anisotropic thermal motion approximation and for the H atoms in the isotropic approximation to R = 0.07. The structure is centrosymmetric and has a threefold axis of pseudosymmetry. The depsipeptide chain is in the form of a bracelet stabilized by six identical intramolecular 4 → 1 hydrogen bonds between the amide C?O and NH groups. The ester carbonyls are oriented towards the symmetry axis, their O atoms forming an ellipsoidal molecular cavity. The isopropyl side chains are located on the molecular periphery. The structure found differs considerably from the conformation of the crystalline naturally occurring antibiotic, valinomycin, but completely resembles that of valinomycin and meso-valinomycin in nonpolar solvents. In the crystal, meso-valinomycin molecules form stacks. The molecular cavities situated in the stacks one above the other along the pseudo-C₃ axis form a continuous channel, the internal surface of which is lined by O atoms. The possible conformations of depsipeptides of the valinomycin series and their mode of action in membranes are discussed in the light of the data obtained.     

Crystal and molecular structure of isoleucinomycin,cyclo[-(D-Ile-Lac-Ile-D-Hyi)3-](C60H102N6O18)

The crystal structure of a synthetic analog of valinomycin, cyclo[-(D -Ile-Lac-Ile-D -Hyi)₃-] (C₆₀H₁₀₂N₆O₁₈), has been determined by x-ray diffraction procedures. The crystals are orthorhombic, space group P2₁2₁2₁, with cell parameters a = 11.516, b = 15.705, c = 39.310 Å, and Z = 4. The atomic coordinates for the C, N, O atoms were refined in the anisotropic thermal motion approximation and for the H atoms in the isotropic approximation. Values of standard (R) and weighted (R_w) reliability factors after refinement are 0.073 and 0.056, respectively. The structure is completely asymmetric. The cyclic molecular backbone is stabilized by six intramolecular hydrogen bonds N? H…?O?C, five bonds being of the 4→1 type and one being of the 5→1 type. The side chains are located on the molecular periphery. The conformational state of isoleucinomycin in the crystal is intermediate between the corresponding crystalline states of valinomycin and meso-valinomycin. The observed conformation suggests that complexation could proceed via entry of the ion at the face possessing the L -Lac residues, the less crowded face.     

Structure of macrocyclic K+, Rb+-complexon of meso-valinomycin monohydrate, cyclo[-(D-Val-Hyi-Val-D-Hyi)3-].H2O, in a crystalline complex with dioxane by x-ray structural data]

The crystal structure of a valinomycin analogue, cyclo[-(D-Val-Hyi-Val-D-Hyi)3-]x(C60H102N6O18) crystallized with dioxane and water molecules, has been solved by X-ray direct methods. The conformation found is analogous to one established for free meso-valinomycin crystallized from other organic solvents. It is characterized by a centrosymmetric bracelet form, stabilized by six intramolecular 4----1 type hydrogen bonds between amide N-H and C = O groups. One water molecule is fixed asymmetrically by hydrogen bonds in the internal negatively charged cavity of the complexon. The meso-valinomycin molecule "bracelets" in the crystal form stacks alternatively with dioxane molecules.     

Crystal and molecular structure of the centrosymmetric meso-valinomycin analogue—cyclo (D-Val-D-Hyi-D-Val-L-Hyi-L-Val-D-Hyi-L-Val-L-Hyi-L-Val-D-Hyi-L-Val-L-Hyi) (C60H102N6O18)

The crystal structure of cyclo (D -Val-D -Hyi-D -Val-L -Hyi-L -Val-D -Hyi-L -Val-L -Hyi-L -Val-D -Hyi-L -Val-L -Hyi)-2H₂O has been solved by x-ray direct methods. The crystals (grown from a mixture of octane / CH₂C₁₂) are an orthorhombic, centrosymmetric space group Pbca, cell parameters a = 11.458(2). b = 25.613(3). c = 23.691(3) Å. Z = 4; therefore the molecule lies on a center of inversion in the cell. The atomic coordinates for the C, N, and O atoms were refined in the anisotropic thermal motion approximation (allowing for II-atom contribution to F_cal) to a standard R- factor value of 0.081. In contrast to meso-valinomycin, the analogue under study does not adopt an octahedral cage bracelet conformation. It has an unusual centrosymmetric elongated form with two type II terminal β-bends formed by N? H?C?O 4 → 1 type intramolecular H bonds. Two symmetry-related water molecules reside in the elongated molecular cavity of the centrosymmetric depsipeptide ring. © 1995 John Wiley & Sons, Inc.     

Crystal and molecular structure of cyclo(L-prolyl-glycyl)3. A cyclic hexapeptide with a cis peptide bond

The crystal structure of cyclo(L-Pro-Gly)3 was solved using X-ray crystallographic techniques. The backbone of the peptide is asymmetric and is made up of five trans peptide units and one cis peptide. There is a hydrogen bonded water bridge that links the carbonyl oxygens, O1 and O4. The molecules exist as dimers in the crystal lattice. The two molecules of the dimer are related by crystallographic twofold symmetry and are linked by two N-H ... O hydrogen bonds. The crystals are trigonal, space group P3(2)12 with a = 11.379(3), c = 32.93(1) and z = 6. The structure was solved by multisolution methods and refined by least squares technique to an R of 0.083.     

Molecular structure of cyclo[-(D-Val-L-Hyi-L-Val-D-Hyi)2-] revealed by x-ray analysis.

The crystal structure of a synthetic analogue of valinomycin, cyclo[-(D-Val-L-Hyi-L-Val-D-Hyi)2-] (octa-meso-valinomycin) (I) (C40H68N4O12.1.5.C4H8O2, M(r) = 937.01 + 88.10), has been determined. Crystals grown from dioxane are monoclinic, space group P2(1)/a, with cell parameters a = 21.487 (8), b = 16.836 (5), c = 16.089 (4) A, beta = 111.70 (4), and Z = 4. The atomic coordinates for nonhydrogen atoms were refined in the anisotropic thermal motion approximation. H atom positions were included in the structure factor calculations at their geometrically expected positions. Values of the standard and weighted R factors after refinement are 0.11 and 0.13, respectively. The conformation of the depsipeptide crystallized from dioxane is different from that crystallized from chloroform (II). The molecule adopts a rectangular shape with two type IV beta-turns containing a hydrogen bond and possesses pseudorotational symmetry. The side chains are located on the molecular periphery. The orientation of the carbonyl groups of the molecule is not conducive for efficient metal-ion coordination and in the observed conformation cannot behave as an ionophore. In the crystal the molecules form infinite chains parallel to the c axis, and are stabilized by two intermolecular hydrogen bonds that are shorter and have better geometry than the intramolecular hydrogen bonds. A phi/psi plot for dodecadepsipeptides with a (DLLD)3 sequence has well-defined areas for Val and Hyi residues only in cases when the crystals have been grown from nonpolar or medium-polar solvents. The phi/psi plot for octadepsipeptides crystallized from chloroform (II) shows this behavior also.(ABSTRACT TRUNCATED AT 250 WORDS)     

Conformation and complexation with metal ions of cyclic hexapeptides: cyclo (L-Leu-L-Phe-L-Pro)2 and cyclo [L-Cys(Acm)-L-Phe-L-Pro]2

Cyclic hexapeptides, cyclo (L-Leu-L-Phe-L-Pro)2 and cyclo[L-Cys(Acm)-L-Phe-L-Pro]2, in which Acm represents an acetoamide-methyl group, were synthesized, and the conformation and complexation with metal ions were investigated. Cooperation of the carbonyl groups of the Cys(Acm) side chains with those of the cyclic skeleton in complexation was especially examined. Cyclo(L-Leu-L-Phe-L-Pro)2, which possesses no functional groups on side chains, was taken as the reference compound. 13C- and two-dimensional n.m.r. measurements revealed that cyclo(L-Leu-L-Phe-L-Pro)2 and cyclo[L-Cys(Acm)-L-Phe-L-Pro]2 took a C2-symmetric conformation containing cis L-Phe-L-Pro bonds in chloroform and acetonitrile. Both cyclic hexapeptides were found to complex selectively with Ba2+ and Ca2+ in acetonitrile. On complexation the conformation of either cyclic hexapeptide changed into a similar one. However, the binding constant of cyclo[L-Cys(Acm)-L-Phe-L-Pro]2 was higher than that of cyclo(L-Leu-L-Phe-L-Pro)2. The n.m.r. measurements showed that the amide carbonyl groups of Cys(Acm) side chains as well as those of cyclic skeleton in cyclo[L-Cys(Acm)-L-Phe-L-Pro]2 cooperatively bound the cations.     

Synthesis and conformation of the cyclic octapeptides cyclo(Phe-Pro)4, cyclo(Leu-Pro)4, and cyclo[Lys(Z)-Pro]4

Cyclic octapeptides, cyclo(X-Pro)₄, where X represents Phe, Leu, or Lys(Z), were synthesized and their conformations investigated. A C₂-symmetric conformer containing two cis peptide bonds was found in all of these cyclic octapeptides. The numbers of available conformations due to the cis–trans isomerization of Pro peptide bonds depended on the nature of the solvent and X residue: they decreased in the following order: cyclo[Lys(Z)-Pro]₄ > cyclo(Leu-Pro)₄ > cyclo(Phe-Pro)₄ in CDCl₃. ¹³C spin-lattice relaxation times (T₁) of these cyclic octapeptides were measured, and the contribution of segmental mobility to T₁ was found to vary with the nature of the X residue.     

Crystal structure and conformation of a cyclic tetrapeptide cyclo(L-Pro-Sar)2 containing all-cis peptide units

The crystal structure and conformation of the synthetic cyclic tetrapeptide, cyclo(L -Pro-Sar)₂, was determined by x-ray analysis. The peptide crystallizes in the orthorhombic space group P2₁2₁2₁ with cell parameters a = 9.277(1), b = 12.884(1), and c = 15.581(2) Å. The crystal structure was solved by the symbolic addition procedure for direct phase determination and least-squares refinement using 1796 reflections, which led to the final R value of 0.043. This structure provides the first example observed in a crystal of a cyclic tetrapeptide in which all four peptide units have been found in the cis conformation with ω angles deviating slightly by 2°–10° from the ideal value of 0°. It was also found that the two Pro C^α-CO single bonds assumed a trans′ (ψ = 159.6° and 158.4°) conformation. Adjoining average planes of the peptide groups fall at nearly right angles to each other. The pyrrolidine ring conformations of the two prolyl residues are in the envelope form, with C^γ carbon out of the least-squares planes for the remaining four atoms.     

The structure of a dinuclear silver(I) complex: Ag2[S2C2(CN)2] [P(C6H5)3]4

The crystal structure of the dimeric Ag maleonitriledithiolate complex, Ag₂[S₂C₂(CN)₂] [P(C₆- H₅)₃]₄ (1), has been performed. Complex 1 crystallizes in the space group P2₁/c with a = 12.2898(77), b = 23.8325(91), c = 23.1790(118) Å, β = 101.315(43)° and Z = 4. Refinement using 3253 reflections with F_o²>3σ(F_o²) yielded R = 0.0662, R_w= 0.0669. The most interesting aspect of the structure is the strong bridging interaction of the chelating maleonitriledithiolate ligand with the second Ag center, where a Ag-S distance of 2.478 Å is observed. The residual bonding capability of the sulfur atoms in the chelating anion [Ag(S₂C₂(CN)₂)(PPh₃)₂]⁻ for [Ag(PPh₃)₂]⁺ is demonstrated.     

Synthesis of [1,2-3H2]cholecalciferol and metabolism of [4-14C,1,2-3H2]- and [4-14C,1-3H]-cholecalciferol in rachitic rats and chicks

[1,2-(3)H(2)]Cholecalciferol has been synthesized with a specific radioactivity of 508mCi/mmol by using tristriphenylphosphinerhodium chloride, the homogeneous hydrogen catalyst. With doses of 125ng (5i.u.) of [4-(14)C,1-(3)H(2)]cholecalciferol the tissue distribution in rachitic rats of cholecalciferol and its metabolites (25-hydroxycholecalciferol and peak P material) was similar to that found in chicken with 500ng doses of the double-labelled vitamin. The only exceptions were rat kidney, with a very high concentration of vitamin D, and rat blood, with a higher proportion of peak P material, containing a substance formed from vitamin D with the loss of hydrogen from C-1. Substance P formed from [4-(14)C,1,2-(3)H(2)]cholecalciferol retained 36% of (3)H, the amount expected from its distribution between C-1 and C-2, the (3)H at C-1 being lost. 25-Hydroxycholecalciferol does not seem to have any specific intracellular localization within the intestine of rachitic chicks. The (3)H-deficient substance P was present in the intestine and bone 1h after a dose of vitamin D and 30min after 25-hydroxycholecalciferol. There was very little 25-hydroxycholecalciferol in intestine at any time-interval, but bone and blood continued to take it up over the 8h experimental period. It is suggested that the intestinal (3)H-deficient substance P originates from outside this tissue. The polar metabolite found in blood and which has retained its (3)H at C-1 is not a precursor of the intestinal (3)H-deficient substance P.     

Studies on cyclic peptides. IV. Conformation of cyclo(Sar-Sar-Gly)2, cyclo(Sar)6 and cyclo(Sar-Gly-Gly)2 and their conformational change induced by alkali thiocyanates

Conformation of cyclo (Sar-Sar-Gly)₂, cyclo(Sar)₆, and cyclo(Sar-Gly-Gly)₂ was investigated by nmr spectroscopy. cyclo(Sar-Sar-Gly)₂, were shown to assume various conformations in dimethysulfoxide. It was attributed to the distribution of cis as well as trans Gly-Sar or Sar-Sar amide links along the peptide backbone. In particular, cyclo(Sar-Sar-Gly)₂ took five or six different conformations: one or three C₂-symmetric conformations and four or three asymmetric conformations, respectively. Three of nine NH resonance signals were ascribed to the internally hydrogen-bonded glycine residues. cyclo(Sar-Sar-Gly)₂ and cyclo(Sar)₆ showed a spectral change on the addition of alkali thiocyanates, indicating a conformational change induced by a complex formation with the alkali cations. The complex nmr spectrum due to a hybridization of different conformations changed with the salt addition into a simple nmr spectrum, suggesting a preponderence of a new, single conformation. On the basis of the spectral change, the strength for the cations binding the cyclic peptides was found to be in the order of K⁺ > Na⁺ > Rb⁺ > Cs⁺ for cyclo(Sar-Gly-Gly)₂ and K⁺ > Rb⁺ > Cs⁺ for cyclo(Sar)₆. On the other hand, cyclo(Sar-Gly-Gly)₂ in dimethylsulfoxide assumed a single C₂ conformation having two glycyl peptide protons shielded from solvent and the other two exposed to solvent. This conformation did not change with the salt addition. Finally, the conformations of several cyclic peptides containing the sarcosine residue such as cyclo(Sar)₆ cyclo(Sar-Sar-Gly)₂ cyclo(Pro-Sar-Gly)₂, and cyclo (Sar-Gly-Gly)₂ were compared. It appeared that proline and glycine residues reduced the conformational multiplicity of the cyclic peptide backbone, and the ability to bind alkali metal cations decreased in the above order.     

Enantioselective synthesis of (R)- and (S)-2-methyl-[3,3,2-2H3] alanines

The synthesis of optically pure (R)- and (S)-2-methyl-[3,3,3-2H3] alanines of biological interest is described. The stereochemistry of the reaction of the lithio derivative of (R)-(-)-2,5-dimethoxy-3-benzyl-3-methyl-3,6-dihydropyrazine with alkyl and deuterated alkyl iodides is discussed. The configuration of the newly formed center of chirality in (R)- and (S)-2-methyl-[3,3,3-2H3] alanines is derived from 1H NMR.     

Metabolism of (1-(13)C) glucose and (2-(13)C, 2-(2)H(3)) acetate in the neuronal and glial compartments of the adult rat brain as detected by [(13)C, (2)H] NMR spectroscopy

Ex vivo ?(13)C, (2)H? NMR spectroscopy allowed to estimate the relative sizes of neuronal and glial glutamate pools and the relative contributions of (1-(13)C) glucose and (2-(13)C, 2-(2)H(3)) acetate to the neuronal and glial tricarboxylic acid cycles of the adult rat brain. Rats were infused during 60 min in the right jugular vein with solutions containing (2-(13)C, 2-(2)H(3)) acetate and (1-(13)C) glucose or (2-(13)C, 2-(2)H(3)) acetate only. At the end of the infusion the brains were frozen in situ and perchloric acid extracts were prepared and analyzed by high resolution (13)C NMR spectroscopy (90.5 MHz). The relative sizes of the neuronal and glial glutamate pools and the contributions of acetyl-CoA molecules derived from (2-(13)C, (2)H(3)) acetate or (1-(13)C) glucose entering the tricarboxylic acid cycles of both compartments, could be determined by the analysis of (2)H-(13)C multiplets and (2)H induced isotopic shifts observed in the C4 carbon resonances of glutamate and glutamine. During the infusions with (2-(13)C, 2-(2)H(3)) acetate and (1-(13)C) glucose, the glial glutamate pool contributed 9% of total cerebral glutamate being derived from (2-(13)C, 2-(2)H(3)) acetyl-CoA (4%), (2-(13)C) acetyl-CoA (3%) and recycled (2-(13)C, 2-(2)H) acetyl-CoA (2%). The neuronal glutamate pool accounted for 91% of the total cerebral glutamate being mainly originated from (2-(13)C) acetyl-CoA (86%) and (2-(13)C, 2-(2)H) acetyl-CoA (5%). During the infusions of (2-(13)C, 2-(2)H(3)) acetate only, the glial glutamate pool contributed 73% of the cerebral glutamate, being derived from (2-(13)C, 2-(2)H(3)) acetyl-CoA (36%), (2-(13)C, 2-(2)H) acetyl-CoA (27%) and (2-(13)C) acetyl-CoA (10%). The neuronal pool contributed 27% of cerebral glutamate being formed from (2-(13)C) acetyl-CoA (11%) and recycled (2-(13)C, 2-(2)H) acetyl-CoA (16%). These results illustrate the potential of ?(13)C, (2)H? NMR spectroscopy as a novel approach to investigate substrate selection and metabolic compartmentation in the adult mammalian brain.     

Solution conformations of two flexible cyclic pentapeptides: cyclo(Gly-Pro-D-Phe-Gly-Ala) and cyclo(Gly-Pro-D-Phe-Gly-Val).

In an effort to explore the residue preferences in three-residue reverse turns (so-called gamma-turns), two cyclic pentapeptides--cyclo(Gly1-Pro2-D-Phe3-Gly4-Ala5) (I) and cyclo(Gly1-Pro2-D-Phe3-Gly4-Val5) (II)--have been synthesized and analyzed by nmr. It was anticipated that the Gly-Pro-D-Phe-Gly portions of these molecules would favor a beta-turn conformation, leaving the remainder of the molecule to adopt a gamma turn, as seen in several previously studied model cyclic pentapeptides. The nmr data for both peptides in CDCl3 (5% DMSO-d6) and in neat DMSO-d6 indicate that the most populated conformation contains a distorted beta turn around Pro2-D-Phe3, which includes a gamma turn around D-Phe3. The distortion in the beta turn does not impede the formation of an inverse gamma turn around residue 5, and indeed, this conformation is observed in both peptides. Both the alanine and the bulkier valine residues are therefore found to be compatible with an inverse gamma turn. Molecular dynamics simulations on the title peptides are reported in the following paper. These simulations indicate that there is conformational flexibility around the D-Phe3-Gly4 peptide bond, which enables the formation of the gamma turn around D-Phe3. The third paper in this series explores the impact of a micellar environment on conformational equilibria in II.     

Lactobacillus plantarum MiLAB 393 produces the antifungal cyclic dipeptides cyclo(L-Phe-L-Pro) and cyclo(L-Phe-trans-4-OH-L-Pro) and 3-phenyllactic acid

We have isolated a Lactobacillus plantarum strain (MiLAB 393) from grass silage that produces broad-spectrum antifungal compounds, active against food- and feed-borne filamentous fungi and yeasts in a dual-culture agar plate assay. Fusarium sporotrichioides and Aspergillus fumigatus were the most sensitive among the molds, and Kluyveromyces marxianus was the most sensitive yeast species. No inhibitory activity could be detected against the mold Penicillium roqueforti or the yeast Zygosaccharomyces bailii. An isolation procedure, employing a microtiter well spore germination bioassay, was devised to isolate active compounds from culture filtrate. Cell-free supernatant was fractionated on a C(18) SPE column, and the 95% aqueous acetonitrile fraction was further separated on a preparative HPLC C(18) column. Fractions active in the bioassay were then fractionated on a porous graphitic carbon column. The structures of the antifungal compounds cyclo(L-Phe-L-Pro), cyclo(L-Phe-trans-4-OH-L-Pro) and 3-phenyllactic acid (L/D isomer ratio, 9:1), were determined by nuclear magnetic resonance spectroscopy, mass spectrometry, and gas chromatography. MIC values against A. fumigatus and P. roqueforti were 20 mg ml(-1) for cyclo(L-Phe-L-Pro) and 7.5 mg ml(-1) for phenyllactic acid. Combinations of the antifungal compounds revealed weak synergistic effects. The production of the antifungal cyclic dipeptides cyclo(L-Phe-L-Pro) and cyclo(L-Phe-trans-4-OH-L-Pro) by lactic acid bacteria is reported here for the first time.     

Complex formation with alkali and alkaline earth metal ions of cyclic octapeptides,cyclo(Phe-Pro)4, cyclo(Leu-Pro)4, and cyclo[Lys(Z)-Pro]4

Complex formation with alkali and alkaline earth metal ions of cyclic octapeptides, cyclo(Phe-Pro)₄, cyclo(Leu-Pro)₄, and cyclo[Lys(Z)-Pro]₄ was investigated in relation to conformation. In an alcohol solution, cyclo(Phe-Pro)₄ did not form complexes. However, cyclo(Leu-Pro)₄ and cyclo[Lys(Z)-Pro]₄ formed complexes selectively with Ba²⁺ and Ca²⁺ ions. Changing the solvent from alcohol to acetonitrile, the complexation behavior was very different. In acetonitrile, cyclo(Phe-Pro)₄ was found to form a complex with Ba²⁺, and CD spectra of cyclo(Leu-Pro)₄ and cyclo[Lys(Z)-Pro]₄ changed sharply on complexation with K⁺. Rate constants of the complex formation between the cyclic octapeptides and metal salts were in the range of 0.7–12 L mol^?1 min^?1 in an alcohol solution. One of the two types of complex formation in acetonitrile was much faster than that in an alcohol solution.     

Synthesis and activity of 2-[4-(4-[3H]-2-cyanophenyl)piperazinyl]-N-(2,4,6-[3H]3-3-methylphenyl)acetamide: a selective dopamine D4 receptor agonist and radioligand

The first selective dopamine D4 agonist radioligand is described. The synthesis of these piperazine radioligands relied on the transformation of brominated precursors 4a and 4b with tritium gas in the presence of a sensitive cyano functional group. The specific activity of these two radioligands was measured and [3H]6b found to be suitable for use in D4 saturation and competition binding studies. The synthesis, biological, and radioactivity of this new agonist radioligand as well as preliminary SAR will be discussed.