Design of Proteinase–Catalyzed Synthesis of Oligopeptides in an Aqueous–Organic Biphasic System

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Thermolysin–catalyzed condensation of N – (benzylxycarbyl) – glycyl – glycyl – L –phenylalanyl–L–leucine ethyl ester (Z–Gly–GlyPheLeuOEt), the precursor of the des–Tyr1–leucine enkephalin from N–(benzyl–oxycarbonyl)–glycyl–glycine (Z–GlyGly) and L–phenylalanyl–L–leucine ethyl ester (PheLeuOEt) was elucidated as a model reaction for the enzymatic synthesis of oligopeptides with many side reactions. In buffer solution, the yield of Z–GlyGly–PheLeuOEt was extremely low with considerable by–products. A large increase in both yield and purity became possible in the aqueous–organic biphasic system by the systematic selection of experimental conditions.

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  1. 1

    Fruton, J.S. 1982. Proteinase-catalyzed synthesis of peptide bonds, p. 239–306. In: Adv. Enzymol., Vol. 53. A. Meister (ed.), John Wiley & Sons, New York.

  2. 2

    Klibanov, A.M., Samokhin, G.P., Martinek, K., and Berezin, I.V. 1977. A new approach to preparative enzymatic synthesis. Biotechnol. Bioeng. 19:1351–1361.

  3. 3

    Martinek, K. and Semenov, A. 1981. Enzymatic synthesis in biphasic aqueous-organic systems II. Shift of ionic equilibria. Biochim. Biophys. Acta 658:90–101.

  4. 4

    Homandberg, G.A., Mattis, J.A., and Laskowski, M. Jr. 1978. Synthesis of peptide bonds by proteinases. Addition of organic co-solvents shifts peptide bond equilibria toward synthesis. Biochemistry 17:5220–5227.

  5. 5

    Lüthi, P. and Luisi, P.L. 1984. Enzymatic synthesis of hydrocarbonsoluble peptides with reverse micelles. J. Am. Chem. Soc. 106:7285–7286.

  6. 6

    Nakanishi, K., Kamikubo, T., and Matsuno, R. 1985. Continuous synthesis of N-(benzyloxycarbonyl)-L-aspartyl-L-phenylalanine methyl ester with immobilized thermolysin in an organic solvent. Bio/Technology 3:459–464.

  7. 7

    Endo, S. 1962. Studies on protease produced by thermophilic bacteria. J. Ferment. Technol. 40:346–353.

  8. 8

    Malfroy, B., Swerts, J.P., Guyon, A., Roques, B.P., and Schwartz, J.C. 1978. High-affinity enkephalin-degrading peptidase in brain is increased after morphine. Nature 276:523–526.

  9. 9

    Zaks, A. and Klibanov, A.M. 1984. Enzymatic catalysis in organic media at 100°C. Science 224:1249–1251.

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Nakanishi, K., Kimura, Y. & Matsuno, R. Design of Proteinase–Catalyzed Synthesis of Oligopeptides in an Aqueous–Organic Biphasic System. Nat Biotechnol 4, 452–454 (1986) doi:10.1038/nbt0586-452

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