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Proceedings of the 31 st European Peptide SymposiumMichal Lebl, Morten Meldal, Knud J. Jensen, Thomas Hoeg-Jensen (Editors)European Peptide Society, 2010New Building Blocks for Solid-Phase Synthesis of PeptideAnalogues: N β -Fmoc-N β -Methyl-aza-β 3 -Amino AcidsKsenija Kisselnova 1,2 , Irène Nicolas 1 , Patrick Bauchat 1 , Jaak Järv 2 ,and Michèle Baudy-Floc’h 11 UMR CNRS 6226, ICMV, University of Rennes 1, Rennes, 35042, France; 2 Institute ofChemistry, University of Tartu, Ravila 14a, Tartu, 50411, EstoniaIntroductionN-Methylation is a precious tool to modify lipophilicity [1], proteolytic stability [2], andbioavailability [3] and to induce conformational rigidity to the peptide backbone [4].However, multiple N-methylation has been seldom employed [5], probably due to theavailability of N-methylated amino acids, as few N-methyl amino acids are commerciallyavailable, and their synthesis is tedious [6].Unnatural aza-β 3 -peptides are synthetic compounds designed to mimic peptides andincrease their bioavailability [7]. They can have chemically diverse side chains, but theyneed to have amide bonds resistant to proteolysis. Yet, additional modifications arerequired to generate peptides with enhanced enzymatic stability and improved oralbioavailability.Results and DiscussionWe have previously reported a method to prepare N β -Fmoc-aza-β 3 -amino acids viareductive amination of glyoxylic acid and Fmoc-hydrazine [8]. Taking into account thisprevious method, we tried to reproduce the same reactions starting from methyl hydrazineR OFmoc NNOHFig. 1. N β -Fmoc-N β -Me-aza-β 3 -aa.instead of hydrazine. Thus the correspondingN-Fmoc-N-Me-hydrazine 2 was prepared using theenhanced nucleophilicity of the methylated N-atomof methylhydrazine 1. Methylhydrazine was treatedat -78°C with fluorenyl chloroformate in thepresence of one equivalent of triethylamine. Themonoacylation occurred on the substituted nitrogenin 92% yield, no acylation was observed on theunsubstituted nitrogen. Then we prepared the N-substituted Fmoc hydrazones bycondensing fluoren-9-yl-methyl carbazate 2 with the appropriate aldehyde or ketone.Reduction of the hydrazone 3 with sodium cyanoborohydride afforded N,N'-trisubstitutedhydrazine 4. From the hydrazine 4 the reductive amination of glyoxylic acid lead to theN β -Fmoc-N β -Methyl-aza-β 3 aa-OH 5 (R = CH 2 R 1 ) (Figure 2 and Table 1).For the monomer glycine a direct reductive amination of glyoxylic acid and fluoren-9-yl-methyl carbazate 2 with NaBH 3 CN and HCl 2N gives the corresponding N β -Fmoc-N β -Methyl aza-β 3 -Gly-OH 5 R=H (Figure 2).Nucleophilic substitution of the corresponding t-butyl bromo acetate with N β -Fmoc-N β -Methyl aza-β 3 -Gly-OH 5 R=H would be an alternative to introduce the side chain forthe aspartic analogue N β -Fmoc-N β -Methyl aza-β 3 -Asp-OH 5 (R=CH 2 CO 2 t-Bu) (Figure 2).MeNHNH 21FmocClNEt 3-78°C92 %FmocNNH 22HOCHCO 2 HR 1 CHOFmoc N R 1 NaBH 3 CN Fmoc N R 1 NaBH 3 CNNN3 HCl 2N4 HCl 2N70-90% 96 %94 %OCHCO 2 HNaBHH3 CN Fmoc NOBrCH 2 CO 2 t-BuHCl 2NN OHDIPEA94 %5 R = H70 %N β -Fmoc-N β -Me-aza-β 3 Gly-OHFmoc NNCO 2 tBuOOHRFmoc NN5 R= CH 2 CO 2 t-BuN β -Fmoc-N β -Me-aza-β 3 Asp-OHOOH5 R= CH 2 R 1N β -Fmoc-N β -Me-aza-β 3 aa-OHFig. 2. Synthesis of N β -Fmoc-N β -Me-aza-β 3 aa 5.104