Product Selectivity in KAHA Ligations: Ester vs. Amide Formation with Cyclic Hydroxylamines

Published as part of the Cluster Recent Advances in Protein and Peptide Synthesis

Abstract

Cyclic hydroxylamines form esters instead of the expected amides as major product upon reaction with α-ketoacids. In this report, we document a systematic investigation into the effect of the hydroxylamine structure and the solvent mixture on the product ratio of amides vs. ester in the KAHA ligation. We show that the ratio is almost exclusively determined by the structure of the hydroxylamine, with only minor contributions from the reaction solvent or the structure of the α-ketoacid.

Key words

ligation - mechanism - hydroxylamines - α-ketoacids - esters - peptides

• References and Notes

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Other amide-forming ligations include the serine/threonine ligation (a) and the traceless Staudinger ligation (b):
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• 3b Kleineweischede R. Hackenberger CP. R. Angew. Chem. Int. Ed. 2008; 47: 5984
• 4 Pattabiraman VR. Ogunkoya AO. Bode JW. Angew. Chem. Int. Ed. 2012; 51: 5114
• 5 Wucherpfennig TG. Rohrbacher F. Pattabiraman VR. Bode JW. Angew. Chem. Int. Ed. 2014; 53: 12244
• 6 Pusterla I. Bode JW. Nat. Chem. 2015; 7: 668
• 7 Rohrbacher F. Deniau G. Luther A. Bode JW. Chem. Sci. 2015; 6: 4889
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• 9 Chéron N. Ramozzi R. Kaïm LE. Grimaud L. Fleurat-Lessard P. J. Org. Chem. 2012; 77: 1361
• 10 General Procedure for KAHA Ligations Ketoacid (1.00–2.00 equiv) and hydroxylamine (1.00–2.00 equiv) were dissolved in 8:2 DMSO/H₂O (20–100 mM) and heated to 60 °C for 12 h. Reaction progress was monitored by analytical HPLC (Shiseido Capcell Pak UG80 C18 column (4.6 × 250 mm), heated to 60 °C, 30–70% MeCN in 20 min). The crude reaction was directly purified by preparative HPLC (Shiseido Capcell Pak MGII C18 column, 20 × 250 mm, r.t., 40–90% MeCN with 0.1% TFA in 20 min, flow rate 10 mL/min). Typical Analytical Data N-(3-Hydroxypropyl)-3-(4-nitrophenyl)propanamide (Amide-24) ¹H NMR (600 MHz, CDCl₃): δ = 8.14–8.10 (m, 2 H, CH), 7.38–7.34 (m, 2 H, CH), 6.07 (br s, 1 H, NH), 3.54 (t, J = 5.6 Hz, 2H, CH₂), 3.37 (q, J = 6.1 Hz, 2 H, CH₂), 3.13 (br s, 1 H, OH), 3.07 (t, J = 7.5 Hz, 2 H, CH₂), 2.53 (t, J = 7.5 Hz, 2 H, CH₂), 1.66–1.59 (m, 2 H, CH₂). ¹³C NMR (150 MHz, CDCl₃): δ = 172.4 (CO), 148.8 (C), 146.7 (CNO₂), 129.4 (CH), 123.9 (CH), 59.6 (CH₂), 37.5 (CH₂), 36.7 (CH₂), 32.1 (CH₂), 31.4 (CH₂). ESI-HRMS: m/z calcd for C₁₂H₁₇N₂O₄ [M + H]⁺: 253.1183; found: 253.1183. 3-{[3-(4-Nitrophenyl)propanoyl]oxy}propan-1-aminium 2,2,2-trifluoroacetate (Ester-24) ¹H NMR (600 MHz, (D₃C)₂SO): δ = 8.19–8.14 (m, 2 H, CH), 7.73 (br s, 3 H, NH₃ ⁺), 7.56–7.51 (m, 2 H, CH), 4.06 (t, J = 6.3 Hz, 2 H, CH₂), 3.00 (t, J = 7.5 Hz, 2 H, CH₂), 2.82 (t, J = 7.4 Hz, 2 H, CH₂), 2.73 (t, J = 7.5 Hz, 2 H, CH₂), 1.88–1.80 (m, 2 H, CH₂). ¹³C NMR (150 MHz, (D₃C)₂SO): δ = 171.9 (CO), 157.9 (q, J = 32.0 Hz, CF₃ COO^–), 148.9 (C), 146.1 (CNO₂), 129.7 (CH), 123.5 (CH), 117.36 (q, J = 301 Hz, CF₃), 61.3 (CH₂), 36.2 (CH₂), 34.1 (CH₂), 29.9 (CH₂), 26.3 (CH₂). ESI-HRMS: m/z calcd for C₁₂H₁₇N₂O₄ [M + H]⁺: 253.1183; found: 253.1186.