Conformational studies of cyclic peptide structures in solution from 1H-Nmr data by distance geometry calculation and restrained energy minimization.

The three-dimensional structure of a cyclic bouvardin analogue, cyclo (-Pro-MeTyr-Ala-MeTyr-MeTyr-D-Ala-) has been determined by distance geometry calculation and restrained energy minimization from nmr data. The preparation of the input for the distance geometry calculations, the modification of the amino acid library, and the analysis of the structures were done with the aid of a recently developed software package, GEOM. A great variety of different initial structures were explored to check the uniqueness of the determined solution structure. Calculations with 500 different initial structures and two different strategies led to a uniquely determined backbone conformation with a root mean square deviations value of 0.4 A. The backbone structure consists of two beta-turns, a beta-II turn at Pro1-MeTyr2, and a beta-VI turn at MeTyr4-MeTyr5. The efficiency of the two calculation strategies were compared in order to propose an optimal means for performing distance geometry calculations with cyclic structures.

2-Epimutalomycin and 28-epimutalomycin, two new polyether antibiotics from Streptomyces mutabilis. Derivatization of mutalomycin and the structure elucidation of two minor metabolites.

A number of derivatives of mutalomycin (1), a naturally occurring polyether antibiotic, have been synthesized. In the desulfurization reaction of the ethylthio derivative (5) of mutalomycin (1) with Raney-nickel we observed an unusual course of the reaction, namely the introduction of a hydroxy group instead of the usual exchange against hydrogen, leading to two reaction products, mutalomycin (1) and 28-epimutalomycin (3). The structure of 3 and 2-epimutalomycin (2), both minor metabolites from the mutalomycin fermentation, were elucidated by X-ray analysis.

A new semisynthetic ergot peptide alkaloid: DCN 203-922.

We report the synthesis, stereochemistry and preliminary pharmacological evaluation of DCN 203-922, a novel ergot alkaloid of the cyclol type, which contains in its peptide moiety the uncommon amino acid L-allo-isoleucine.

Conformational analysis of two somatostatin analogues by 1H 500 MHz n.m.r. spectroscopy.

A series of nine closely related somatostatin analogues, containing the hexapeptide H-Cys2-Phe3-D-Trp4-Lys5-Thr6-Cys7-NH2 sequence have been synthesized by Bauer et al. The conformational properties of two of them, showing intermediate activities between those of SMS 201-995 and somatostatin, have been studied by high field n.m.r. spectroscopy in DMSO. Assignments were made using 2D-n.m.r. methods, in particular NOESY experiments and detection of long-range connectivities in aromatic residues. In all the compounds of this series, the biologically active ones as well as the inactive ones, the n.m.r. parameters are in favour of a predominant conformation with a type II' beta turn involving amino acids Phe3 to Thr6. A clearcut correlation exists between the predominant conformation at the cystine bridge side and the activity. The presence of the exocyclic amino acids Phe1 and Thr8 (ol) plays a major role in stabilization of the active conformation.

Clavamycins, new clavam antibiotics from two variants of Streptomyces hygroscopicus. II. Isolation and structures of clavamycins A, B and C from Streptomyces hygroscopicus NRRL 15846, and of clavamycins D, E and F from Streptomyces hygroscopicus NRRL 15879.

Clavamycins A, B, C, D, E and F represent new members of the clavam antibiotics group. Their purification was achieved by a variety of preparative chromatographic methods, essentially on reversed phase carrier. The structures were deduced from spectroscopic properties, especially from extensive NMR studies.

SMS 201-995, a very potent analogue of somatostatin. Assignment of the 1H 500 MHz n.m.r. spectra and conformational analysis in aqueous solution.

The conformations of a cyclic analogue of somatostatin, SMS 201-995, have been studied by n.m.r. spectroscopy at 500 MHz in aqueous solution. Assignments were made by use of 2D-correlated methods, especially by detecting long-range connectivities in order to identify the aromic amino-acid and long-range couplings between alpha protons of consecutive residues. Measurements of temperature coefficients of amide protons and of NH-C alpha H coupling constants enabled us to conclude that in water the molecule is rather flexible, with no evidence for a beta turn structure involving Thr6. An equilibrium involving two gamma turn conformations stabilized respectively by Cys2-D-Trp4 and Phe3-Lys5 hydrogen bonds, is responsible for the large upfield shift observed for the Lys5 gamma protons and is compatible with the measured JNH-C alpha H coupling constants.

SMS 201-995, an octapeptide somatostatin analogue. Assignment of the 1H 500 MHZ n.m.r. spectra and conformational analysis of SMS 201-995 in dimethylsulfoxide.

The possible conformations of SMS 201-995, an active analogue of somastostatin, have been studied in dimethylsulfoxide solution by 500 MHz proton n.m.r. spectroscopy. The assignments have been made by use of 2D-correlated methods to detect long-range coupling connectivities in aromatic residues and between the alpha protons of consecutive residues. NOESY experiments enabled us to correlate amide and alpha protons of neighbouring amino acid residues, which indicate a less flexible situation than in water. Measurements of temperature coefficients of the amide protons, of NH-C alpha H coupling constants and NOE effects are in favour of one predominant conformation with a beta turn, of type II', involving amino acids Phe3 to Thr6.

Hexapeptide analogue of somatostatin, Sandoz 201-456. Conformational study in dimethylsulfoxide by 1D and 2D n.m.r. methods.

The conformational properties of the somatostatin analogue 201-456 (1) have been studied by high field n.m.r. in DMSO. This analogue is the base structure of nine derivates synthesized by Bauer et al. and shows a very low biological activity, although derived structures such as SMS 201-995 (2) are very potent. Our study has shown an important difference between the most stable conformation of the two compounds: although the beta turn type II' structure at the Phe3-Trp4-Lys5 level is present in both analogues, an important conformational change appears at the cystine bridge. In SMS 201-995 the beta turn/beta sheet conformation is stabilized by the additional amino-acids D-Phe1 and Thr8 (ol) through intramolecular H-bonds.

Disposition of cyclosporine in several animal species and man. I. Structural elucidation of its metabolites.

Nine ether-extractable metabolites of cyclosporine were isolated from urine of dog and man and from rat bile and feces and purified by preparative HPLC and TLC. Structural assignments were mainly based on spectroscopic data (1H NMR, 13C NMR, MS) and the results of the amino acid analysis after hydrolysis with hydrochloric acid. All the identified metabolites retained the intact cyclic oligopeptide structure of the parent drug. Structural modifications originated from enzymatic oxidation at specific sites of the peptide subunits. Transformation processes principally involved hydroxylation at the terminal carbon atom (eta-position) of the C9-amino acid 1 and the gamma-position of the N-methylleucines 4, 6, and 9, as well as N-demethylation of the N-methylleucine 4. Regioisomeric monohydroxylated cyclosporines (metabolites 1, and 17) and N-demethylcyclosporine (metabolite 21) were the primary metabolites resulting from hydroxylation of the C9-amino-acid 1 and the N-methylleucine 9, and from N-demethylation of the N-methylleucine 4. Dihydroxylated derivatives of cyclosporine (metabolites 8, 10, and 16) were generated by further oxidation of metabolite 1 on one of the other N-methylleucines (4 or 6) or on the C9-amino acid 1, or of metabolite 17 on the N-methylleucine 9. More extensive modifications were observed for metabolite 9, a dihydroxy-N-demethylcyclosporine, which could have been formed from the dihydroxy derivative 16 by N-demethylation or from the N-demethylcyclosporine 21 by dihydroxylation. Metabolite 18 differed from 17 (monohydroxycyclosporine) by the presence of a cyclic ether moiety, formally derived by intramolecular addition of the beta-hydroxyl group to the double bond of the C9-amino acid 1.(ABSTRACT TRUNCATED AT 250 WORDS)

Contribution to knowledge of the biosynthesis of cyclosporin A.

3H- and 13C-NMR spectroscopic investigations on the structure of labeled cyclosporin A were performed after feeding of appropriate precursors. The 6 N-methyl groups and the methyl group in position 4 of the epsilon, zeta-unsaturated amino acid No. 1 (Mebmt) are introduced as intact CH3-units from methionine. Four head-to-tail acetate units are involved in the biosynthesis of the 8-carbon chain of the olefinic amino acid.

Fate and disposition of bromocriptine in animals and man. I: structure elucidation of the metabolites.

A total of 16 metabolites of bromocriptine could be isolated from rat bile and incubates of rat liver cell preparations using [6-methyl-14C]bromocriptine as substrate. Separation and purification was achieved by reversed-phase liquid chromatography and preparative thin-layer chromatography in conjunction with radioactivity monitoring. Structure elucidation was based on spectroscopic data (UV, IR, NMR, EI- and FD-MS) and the results of amino acid analysis after acid hydrolysis. Based on the identified metabolites four principal transformation process could be described: -Hydrolytic cleavage of the amide bridge leading to the formation of 2-bromolysergic acid amide (3) and 2-bromolysergic acid (7). -epimerization at position 8 of the bromolysergic acid moiety to the iso-derivatives (isobromocriptine, 2-bromo-isolysergic acid (6), its amide (1), etc.) -regiospecific oxidation at position 8' in the proline fragment generating stereoisomeric 8'-hydroxy-bromocriptines (21-24) -further oxidation of the 8'-hydroxylated derivatives by either the introduction of a second hydroxy group at position 9' to give dihydroxylated derivatives (detected as conjugates with glucuronic acid: metabolites 29, 30 and 31), or the opening of the proline ring under formation of the metabolites 4 and 5 containing glutamic acid instead of proline (7', 8'-seco- 8'-carboxy-bromocriptines). It is suggested that the primary and principal metabolic attack occurs at the proline fragment of the drug. In contrast to the biotransformation of ergoline compounds, none of the bromocriptine metabolites detected showed oxidative transformations in the lysergic acid half of the molecule.

Tetronomycin, a novel polyether of unusual structure.

Tetronomycin, C34H50O8, isolated from a strain of Streptomyces sp. nov. represents a novel polycyclic ionophore polyether. The crystal structure and absolute configuration were established by X-ray analysis of the mono-O-acetyltetronomycin silver salt. Tetronomycin is the first metabolic polyether which contains a tetronic acid moiety instead of the essential carboxylic acid function. A trisubstituted cyclohexane ring and an interesting molecular conformation of the silver salt represent additional unique structural features. Extensive NMR-studies enabled the assignment of chemical shifts and the correlation of the proton and carbon signals. Tetronomycin exhibits activity against Gram-positive bacteria.

Noboritomycins A and B, new polyether antibiotics.

Noboritomycins A and B, two new polycyclic ionophoric polyethers were isolated from a strain of Streptomyces noboritoensis. The crystal structure and absolute configuration of noboritomycin A were established by X-ray analysis of its silver salt C43/63O14Ag. Noboritomycin A is the first metabolic polyether possessing two carboxylic acid functions on the carbon backbone (C-31), namely a free acid and an additional carboxylic acid ethylester group. An unusual spiroketal system as well as a salicylic acid chromophore represent further remarkable elements. Noboritomycin A shows in this respect a structural relationship to salinomycin and lasalocid respectively. Comparison of physico-chemical data, in particular the interpretation of the 1H- and 13C-NMR spectra, revealed that noboritomycins A and B are structurally closely related, noboritomycin B carrying an ethyl substituent on the aromatic ring in the place of a methyl group present in noboritomycin A. Both metabolites exhibit activity against Gram-positive bacteria and against Eimeria tenella (chicken coccidiosis).