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ABSTRACT: In this paper we report the preparation of specifically 2H-labelled L-glutamic acids and 2H-, 15N-, 13C-labelled L-glutamines on the gram scale. The products obtained have high isotope enrichment (>98%) and high optical purity. The synthetic schemes allow the specific isotope enrichment of every H, C, and N position or any combination of positions. (2-2H)-L-Glutamic acid was synthesized by enantioselective enzymatic conversion of 2-oxoglutaric acid using glutamate dehydrogenase (GDH, E.C. 1.4.1.3.), alcohol dehydrogenase (ADH, E.C. 1.1.1.1.) and (2H6)ethanol. (3,3-2H2)-L-Glutamic acid was prepared by enzymatic conversion of 2-oxo-3,3-di-deuteroglutaric acid which was easily obtained from 2-oxoglutaric acid by an isotope exchange reaction in 2H2O at pH 13.0. (4,4-2H2)-L-Glutamic acid was obtained by chemical exchange in 20% 2HCl. Four different isotopomers of L-glutamine [(2-2H)-, (3,3-2H2)-, (4,4-2H2)-, and (5-13C)-L-Gln] were synthesized by the enantioselective conversion of isotopically labelled L-glutamic acids using glutamine synthetase (GS, E.C. 6.3.1.2.). The amide group of glutamine was labelled with 15N using 15NH4Cl in the enzymatic reaction. The labelled L-glutamic acids and L-glutamines were characterized by 1H-NMR, 13C-NMR and mass spectrometry.
Recueil des Travaux Chimiques des Pays-Bas. 09/2010; 113(7‐8):369 - 375.
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ABSTRACT: The topologies of zervamicin II and alamethicin, labeled with (15)N uniformly, selectively, or specifically, have been investigated by oriented proton-decoupled (15)N solid-state NMR spectroscopy. Whereas at lipid-to-peptide (L/P) ratios of 50 (wt/wt) zervamicin II exhibits transmembrane alignments in 1,2-dicapryl (di-C10:0-PC) and 1,2-dilauroyl (di-C12:0-PC) phosphatidylcholine bilayers, it adopts orientations predominantly parallel to the membrane surface when the lengths of the fatty acyl chains are extended. The orientational order of zervamicin II increases with higher phospholipid concentrations, and considerable line narrowing is obtained in di-C10:0-PC/zervamicin II membranes at L/P ratios of 100 (wt/wt). In contrast to zervamicin, alamethicin is transmembrane throughout most, if not all, of its length when reconstituted into 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine bilayers. The (31)P solid-state NMR spectra of all phospholipid/peptaibol samples investigated show a high degree of headgroup order, indicating that the peptides do not distort the bilayer structure. The observed differences in peptide orientation between zervamicin and alamethicin are discussed with reference to differences in their lengths, helical conformations, distribution of (hydroxy)proline residues, and hydrophobic moments. Possible implications for peptaibol voltage-gating are also described.
Biochemistry 09/2001; 40(31):9428-37. · 3.42 Impact Factor
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ABSTRACT: Approach was developed to a preparative synthesis of isotope-labeled aminoacids contained in servamycin IIB antibiotic. Glutamines labeled with 15N, 13N, and 2H were prepared in 70-80% yield starting with the corresponding labeled glutamic acids under catalysis with the glutamine synthetase enzyme. 1
5N-2-aminoisobutanoic acid and 15N-isovaline were obtained by Strecker method in 65 and 31% yields respectively. All compounds synthesized were identified and characterized by NMR spectroscopy.
Russian Journal of Organic Chemistry 03/2001; 37(4):475-479. · 0.65 Impact Factor
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ABSTRACT: For the first time the total synthesis of the peptaibol zervamicin IIB is described. Synthesis of this peptaibol was achieved by the Fmoc/tert-butyl strategy in solution using a fragment condensation approach. Three fragments of zervamicin IIB were obtained by stepwise elongation with Fmoc amino acids using BOP as a coupling reagent. For the introduction of the highly sterically hindered alpha-aminoisobutyric acid residues BOP/DMAP activation was applied. The fmoc group was removed by reaction with 0.1 M NaOH in dioxane/methanol/water (30/9/1, v/v/v). Peptide fragments were coupled by means of a new coupling reagent, CF3-PyBOP. Using the strategy developed, zervamicin IIB and two analogues specifically deuterium-labelled at different positions of the glutamine-11 residue have been synthesized in 40% overall yield based on the isotopically labelled amino acid and with 98 +/- 2% of isotope enrichment. FAB mass spectroscopy, 600 MHz 1H-NMR spectroscopy and high-performance liquid chromatography provided convincing evidence that the synthetic products, zervamicin IIB and its deuterium-labelled analogues, fully correspond to the naturally occurring zervamicin IIB.
Journal of Peptide Science 3(3):193-208. · 1.80 Impact Factor
