The Goldschmidt-Wick type reaction between isocyanates of NÎ±-Fmoc-amino acids/peptide acids and NÎ±-Boc-/Z-/Bsmoc-amino acids catalyzed by DMAP leads to the incorporation of a reversed peptide bond. It was found to be a simple, efficient and clean reaction. All the retro-inverso peptides made were obtained as crystalline compounds in 70-92 yields. Â© 2006 Elsevier Ltd. All rights reserved

Sureshbabu, V.V.

Venkataramanarao, R.

ePrints@Bangalore University

Tetrahedron Letters 47 (2006) 9139–9141Synthesis of retro-inverso peptides employing isocyanates ofNa-Fmoc-amino acids/peptide acids catalyzed by DMAPIRao Venkataramanarao and Vommina V. Sureshbabu*Department of Studies in Chemistry, Central College Campus, Bangalore University, Dr. B.R. Ambedkar Veedhi,Bangalore 560 001, IndiaReceived 14 June 2006; revised 27 September 2006; accepted 12 October 2006Available online 2 November 2006Abstract—The Goldschmidt–Wick type reaction between isocyanates of Na-Fmoc-amino acids/peptide acids and Na-Boc-/Z-/Bsmoc-amino acids catalyzed by DMAP leads to the incorporation of a reversed peptide bond. It was found to be a simple, eﬃcientand clean reaction. All the retro-inverso peptides made were obtained as crystalline compounds in 70–92% yields. 2006 Elsevier Ltd. All rights reserved.The concept of retro-inverso peptides with free andblocked C- and N-termini has led to new molecules withimproved biological activity based on conformationaland topological properties.1 Important classes of mole-cules studied include neurotransmitters, hormones,inhibitors of proteases and protein kinases, sweeteners,antimicrobial peptides, adhesion molecules, antigenicepitopes, immunomodulators and immunological probes.2A large number of molecules have been synthesized bytwo approaches.3,4 The ﬁrst protocol developed by Cho-rev et al. involves the preferential use of acetyl protectinggroups rather than carbamates (speciﬁcally Boc or Zmoi-eties) due to the stability of isocyanate, which avoids thedisplacement reactions.5,6OHFmoc-NHR1ON3Fmoc-NHR1O+aScheme 1. Reagents and conditions: (a) NMM, IBC-Cl, 0 C, aq NaN3, 30 m(ii) Boc-/Z-amino acid, HBTU, DIEA.0040-4039/$ - see front matter  2006 Elsevier Ltd. All rights reserved.doi:10.1016/j.tetlet.2006.10.066qPresented at the 42nd Japanese Peptide Society, Osaka, Japan, held in 200* Corresponding author. Tel.: +91 0822961339; e-mail: hariccb@rediﬀmail.cThe more commonly used route involves a Hofmannrearrangement of Na-protected amino acid amides usingiodobenzene bis(triﬂuoroacetate) (IBTFA) as an oxidiz-ing agent to obtain mono N-acetylated, gem-diamino-alkyl triﬂuoroacetates as key intermediates.7–9 Asconcluded recently by Chorev, rigorous puriﬁcation isrequired for building blocks; thus selection of protectinggroups and the presence of reactive side-chains are keyfactors which cannot be overlooked.3 Further, IBTFAoxidation is not compatible with Fmoc chemistry.10Acid azides11,12 and isocyanates13 derived from Fmocprotected natural amino acids can be prepared easilyand can also be isolated, characterized and employedas key intermediates at ambient temperature in the syn-PhCH2OH Fmoc-NHR1NH-ZHNFmoc-R1NHOR2NH-Boc/Zbc1in; (b) microwave irradiation for four 15 min intervals; (c) (i) Pd/C, H2,5.omHN NCOCOOH Fmoc-R1R1HNFmoc-R1NH-Bsmoc-/Z-/BocNHOR2Fmoc-HNa b2Scheme 2. Reagents and conditions: (a) NMM, IBC-Cl, 0 C, aq NaN3, 30 min, microwave irradiation in toluene, 1 min or reﬂux 1 h; (b) Boc-/Z-/Bsmoc-amino acid, cat. DMAP, temp 0 C to rt.HNOOHOHNOHNNH-Boc/ZO2 23Fmoc-NHa, bFmoc-NHR1R2R3R1R2Scheme 3. Reagents and conditions: (a) NMM, IBC-Cl, 0 C, aq NaN3, 30 min, microwave irradiation in toluene, 1 min or reﬂux for 1 h; (b) Boc-/Z-amino acid, cat. DMAP, temp 0 C to rt.Table 1. Retro-inverso peptides synthesized through the DMAP catalyzed methodEntry Compound Mp (C) Yield(%)Mass 1H NMRa Fmoc-gLeu-rPhe-Boc 174 90 594.7032 d 0.93 (d, J = 5.1 Hz, 6H), 1.33 (m, 2H), 1.37 (s, 9H), 1.62 (m, 1H), 3.10 (d,J = 6.2 Hz, 2H), 3.77 (m, 1H), 3.96 (m, 1H), 4.19 (t, J = 6.6 Hz, 1H), 4.42(d, J = 5.9 Hz, 2H), 5.08 (br s, 1H), 5.62 (br s, 1H), 6.20 (br s, 1H), 7.17–7.77 (m, 13H)b Fmoc-gAla-rLeu-Boc 190 92 518.2631 0.82 (m, J = 5.2 Hz, 6H), 1.21 (d, J = 6.2 Hz, 3H), 1.34 (s, 9H), 1.42 (m,2H), 4.18 (t, J = 6.6 Hz, 1H), 4.21 (d, J = 6.5 Hz, 2H), 5.10 (d, J = 8.8 Hz,1H), 6.10 (d, J = 6.6 Hz, 1H), 6.35 (d, J = 9.2 Hz, 1H), 7.31–7.87 (m, 8H)c Fmoc-gIle-rCys(Acm)-Boc 202 75 621.7504 0.91 (m, 6H), 1.33 (s, 9H), 1.41 (m, 1H), 1.56 (m, 2H), 2.30 (s, 3H), 3.11 (d,J = 6.2 Hz, 2H), 3.62 (m, 1H), 4.19 (t, J = 6.6 Hz, 1H), 4.42 (d, J = 6.5 Hz,2H), 4.27 (m, 1H), 5.12 (br d, J = 9.2 Hz, 2H), 7.27–7.77 (m, 13H)d Fmoc-gLeu-rAsp(Bzl)-Boc 198 79 652.7466 0.82 (m, J = 5.1 Hz, 6H), 1.34 (s, 9H), 1.42 (m, 2H), 2.21–2.56 (m, 1H),3.00–3.33 (m, 1H), 4.21 (t, J = 6.6 Hz, 1H), 4.42 (d, J = 6.9 Hz, 2H), 5.11(m, 2H), 5.21 (d, J = 8.8 Hz, 1H), 6.20 (d, J = 6.6 Hz, 1H), 6.38 (d,J = 9.2 Hz, 1H), 7.24–7.87 (m, 13H)e Fmoc-gPhe-rVal-Bsmoc 172 89 702.7854 0.94 (t, J = 13.2 Hz, 6H), 1.89 (m, 1H), 3.19 (d, J = 6.1 Hz, 2H), 3.91 (br s,1H), 4.19 (t, J = 6.6 Hz, 1H), 4.25 (br s, 1H), 4.42 (d, J = 6.5 Hz, 2H), 5.21(s, 2H), 7.13–7.79 (m, 18H)f Fmoc-gAla-rPhe-Bsmoc 180 85 674.7295 1.30 (d, J = 6.3 Hz, 3H), 2.96 (d, J = 6.0 Hz, 2H), 3.33 (m, 1H), 3.88 (m,1H), 4.19 (t, J = 6.6 Hz, 1H), 4.42 (d, J = 6.9 Hz, 2H), 5.21 (d, J = 8.8 Hz,2H), 5.91 (br s, 1H), 6.71 (br s, 1H), 7.13–7.90 (m, 17H)g Fmoc-gAla-rVal-Boc 218 88 504.5642 0.94 (t, J = 13.2 Hz, 6H), 1.33 (s, 9H), 1.39 (d, J = 6.4 Hz, 3H), 1.89 (m,1H), 3.87 (br s, 1H), 4.19 (t, J = 6.6 Hz, 1H), 4.26 (br s, 1H), 4.42 (d,J = 6.5 Hz, 2H), 7.39–7.78 (m, 8H)h Fmoc-gGly-rPhe-Boc 166 85 538.6246 1.33 (s, 9H), 3.21 (d, J = 4.4 Hz, 1H), 3.88 (br s, 1H), 4.19 (t, J = 6.6 Hz,1H), 4.30 (br d, J = 9.2 Hz, 2H), 4.42 (d, J = 6.5 Hz, 2H), 7.13–7.79 (m,13H)i Fmoc-gSer(tBu)-rLeu-Z 171 87 624.7435 0.92 (d, J = 5.2 Hz, 6H), 1.33 (m, 2H), 1.38 (s, 9H), 1.62 (m, 1H), 3.55 (brm, 2H), 3.91 (br s, 1H), 4.10 (m, 1H), 4.19 (t, J = 6.6 Hz, 1H), 4.42 (d,J = 6.9 Hz, 2H), 5.10 (s, 2H), 7.13–7.79 (m, 13H)j Fmoc-Phe-gAla-rLeu-Boc 110 88 665.7854 0.90 (d, J = 5.4 Hz, 6H), 1.33 (m, J = 6.0 Hz, 2H), 1.37 (s, 9H), 1.62 (m,1H), 3.19 (d, J = 6.2 Hz, 2H), 3.88 (m, 1H), 4.10 (m, 1H), 4.19 (t, J =6.6 Hz, 1H), 4.42 (d, J = 6.9 Hz, 2H), 7.13–7.79 (m, 13H)k Fmoc-Leu-gPhe-rPro-Boc 105 91 691.3472 0.88 (d, J = 5.4 Hz, 6H), 1.25 (m, 2H), 1.41 (s, 9H), 1.55 (m, 1H), 1.78 (m,1H), 1.97 (m, 4H), 3.18 (d, J = 6.2 Hz, 2H), 3.35 (m, 2H), 3.88 (br s, 1H),4.19 (t, J = 6.6 Hz, 1H), 4.42 (d, J = 6.9 Hz, 2H), 7.13–7.79 (m, 13H)l Fmoc-Val-gAla-rPhe-Z 123 89 685.7735 0.93 (t, J = 13.2 Hz, 6H), 1.39 (d, J = 6.6 Hz, 3H), 1.89 (m, 1H), 3.20 (m,2H), 3.81 (br s, 1H), 4.19 (t, J = 6.6 Hz, 1H), 4.42 (d, J = 6.9 Hz, 2H), 5.12(d, J = 9.2 Hz, 2H), 7.21–7.81 (m, 18H)m Fmoc-Ala-gLeu-rAib-Z 134 82 637.7371 0.92 (d, J = 5.2 Hz, 6H), 1.32 (m, J = 6.4 Hz, 2H), 1.39 (d, J = 6.6 Hz, 3H),1.43 (s, 6H), 3.90 (m, 1H), 4.19 (t, J = 6.6 Hz, 1H), 4.42 (d, J = 6.5 Hz, 2H),5.12 (d, J = 9.2 Hz, 2H), 7.21–7.81 (m, 13H)9140 R. Venkataramanarao, V. V. Sureshbabu / Tetrahedron Letters 47 (2006) 9139–9141R. Venkataramanarao, V. V. Sureshbabu / Tetrahedron Letters 47 (2006) 9139–9141 9141thesis of peptides, peptidomimetics and various methylcarbamates.14 This letter demonstrates the utility ofisocyanates derived from Fmoc-a-amino acids for thesynthesis of retro-inverso peptides by the Goldsch-midt–Wick type reaction.In the context of our ongoing interest in the incorpora-tion of retro-inverso bonds in peptides, we initiallyenvisaged a route involving the preparation of N,N 0-bis-protected gem-diamines (Scheme 1). After deprotectionof the Z group employing Pd/C, the coupling of Boc-/Z-amino acids with Fmoc–NH–CHR1–NH2 usingHBTU was explored. In addition to the multi-stepprotocol, the alcoholysis of isocyanates has to be carriedout under microwave irradiation for about 15 min.Recently, Gurtler et al. developed a Mg catalyst systemfor the reaction of aliphatic isocyanates and blockedisocyanates with carboxylic acids. However, our eﬀortsto couple the isocyanate of Fmoc-Leu-OH with Boc-Phe-OH in the presence of Mg as a catalyst wasunsuccessful.15Later we found that a catalytic amount of DMAP16,17accelerates the coupling leading to the formation of aretro-inverso peptide bond. In a typical reaction,Fmoc-amino acid azides were prepared by generatinga mixed anhydride of Fmoc-amino acid and then reac-tion with NaN3. The resulting azide was dissolved in tol-uene and subjected to Curtius rearrangement.18 Afterevaporation of toluene under reduced pressure, it wasdissolved in DCM and a mixture of Boc-/Z-/Bsmoc-amino acid and a catalytic amount of DMAP wereadded at 0 C and the mixture was stirred for 30 min,allowed to come to rt and stirring was continued foranother 2 h. A simple work-up and recrystallizationled to the isolation of product 2 in 70–92% yield(Scheme 2). The same methodology has also been ap-plied to Fmoc-peptide acids which resulted in products3 (Scheme 3).19 The entire course of the reaction canbe completed in about 4 h. All the retro-inverso peptidesmade were isolated by a single recrystallization (Table 1)and were fully characterized by 1H NMR, 13C NMRand mass spectroscopic measurements.In conclusion, the Goldschmidt–Wick type reactionbetween isocyanates of Fmoc-amino acids and Na-protected amino acids catalyzed by DMAP results inretro-inverso peptides. The protocol is simple, eﬃcientand scale-up of the reaction up to 25 mmol per batchdid not pose any problems. Thus, it is now demon-strated that the Fmoc group for Na-protection duringthe synthesis of retro-inverso peptides permits the useof a urethane as a protecting group which can be easilydeprotected, unlike an N-acetyl, and can also allowfurther extension of the peptide chain.AcknowledgementsThe authors thank Professor Fred Naider, The Collegeof Staten Island, City University of New York, NewYork for useful discussions. We also thank the Depart-ment of Biotechnology, Government of India for theﬁnancial assistance. R. V. R. Rao thanks the CSIR,New Delhi, for the award of a fellowship.References and notes1. Chorev, M. Biopolymers 2005, 80, 67–84.2. Chorev, M.; Goodman, M. Acc. Chem. Res. 1993, 26,266–273.3. Scheibler, L.; Chorev, M. Methods of Organic Chemistry,Synthesis of Peptides and Peptidomimetics; Houben-Weyl:New York, 2002, E 22c, pp 528–550.4. For other protocols for the synthesis of retro-inversopeptides, see Ref. 3.5. Chorev, M.; MacDonald, S. A.; Goodman, M. J. Org.Chem. 1984, 49, 821–827.6. Fletcher, M. D.; Campbell, M. M. Chem. Rev. 1998, 98,763–795.7. Sisto, A.; Verdini, A. S.; Virdia, A. Synthesis 1985, 294–296.8. Boutin, R. H.; Loudon, G. M. J. Org. Chem. 1984, 49,4277–4284.9. Yamamoto, Y.; Yatagai, H.; Maruyama, K. J. Org. Chem.1979, 44, 1746–1747.10. Englund, E. A.; Gopi, H. N.; Appella, D. H. Org. Lett.2004, 6, 213–215.11. Suresh Babu, V. V.; Ananda, K.; Vasanthakumar, G. R.J. Chem. Soc., Perkin. Trans. 1 2000, 4328–4331.12. Vasanthakumar, G. R.; Ananda, K.; Sureshbabu, V. V.Indian J. Chem. 2002, 41B, 1733–1735.13. Patil, B. S.; Vasanthakumar, G. R.; Suresh Babu, V. V. J.Org. Chem. 2003, 68, 7274–7280.14. Patil, B. S.; Suresh Babu, V. V. Lett. Pept. Sci. 2004, 10,93–97.15. (a) Gurtler, C.; Danielmeier, K. Tetrahedron Lett. 2004,45, 2515–2521; (b) Gertzmann, R.; Gurtler, C. Tetra-hedron Lett. 2005, 46, 6659–6662.16. Schuemacher, A. C.; Hoﬀmann, R. W. Synthesis 2001,243–246.17. Ramiah, M.; Scriven, E. F. V. Aldrichim. Acta 2003, 36,21–27.18. General procedure for the synthesis of isocyanates of Na-Fmoc-amino acids/peptide acids: To an ice-cold solutionof Na-Fmoc-amino acid or peptide acid (1 mmol) in dryTHF (5 mL) were added N-methylmorpholine (NMM)(0.11 mL, 1 mmol) and IBC-Cl (0.135 mL, 1 mmol) andthe mixture was stirred at 10 C for 5 min. The resultingreaction mixture was treated with aq NaN3 (0.098 g,1.5 mmol in 1 mL) and stirred for another 30 min. Aftercompletion of the reaction, the organic layer was evapo-rated and the residue was dissolved in CH2Cl2 (30 mL),washed with 10 mL each of 5% HCl, 5% aq NaHCO3 andbrine and dried over anhydrous Na2SO4. The solvent wasevaporated in vacuo and the residue dissolved in toluene(10 mL) and heated at 90 C under nitrogen. After 1 h, thesolvent was removed under reduced pressure to give theisocyanate, which was used for the next step.19. General procedure for the synthesis of retro-inverso pep-tides: To a mixture of Na-Fmoc-amino acid/peptideisocyanate (1 mmol) and Boc-/Z-amino acid (1.2 mmol)in dry CH2Cl2 (20 mL) at 0 C was added DMAP(0.3 mmol). After 30 min at 0 C, stirring was continuedat rt for another 2 h. The solvent was evaporated in vacuoand the residue was dissolved in ethyl acetate, washed with10 mL each of 5% Na2CO3, 5% citric acid and brine anddried over anhydrous Na2SO4. The solvent was evapo-rated and the resulting residue crystallized using ethylacetate/hexane (2:8).

Synthesis of retro-inverso peptides employing isocyanates of NÎ±-Fmoc-amino acids/peptide acids catalyzed by DMAP

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Synthesis of retro-inverso peptides employing isocyanates of NÎ±-Fmoc-amino acids/peptide acids catalyzed by DMAP

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