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Amino Acids

, Volume 17, Issue 3, pp 285–291 | Cite as

Trypsin-catalyzed peptide synthesis withm-guanidinophenyl andm-(guanidinomethyl)phenyl esters as acyl donor component

  • H. Sekizaki
  • K. Itoh
  • E. Toyota
  • K. Tanizawa
Full Papers

Summary

Two series of inverse substrates,m-guanidinophenyl andm-(guanidinomethyl)phenyl esters derived fromN-(tert-butyloxycarbonyl)amino acid, were prepared as an acyl donor component for trypsin-catalyzed peptide synthesis. The kinetic behavior of these esters toward tryptic hydrolysis was analyzed. They were found to couple with an acyl acceptor such asl-alaninep-nitroanilide to produce dipeptide in the presence of trypsin.Streptomyces griseus trypsin was a more efficient catalyst than the bovine trypsin. Within the enzymatic peptide coupling methods, this approach was shown to be advantageous, since the resulting peptides are resistant to the enzymatic hydrolysis.

Keywords

Amino acid esters Inverse substrate Kinetics of tryptic hydrolysis Protease catalysis Peptide synthesis 

Abbreviations

Boc

tert-butyloxycarbonyl

Aib

α-aminoisobutyric acid

DMSO

dimethylsulfoxide

Tris

tris(hydroxymethyl)aminomethane

MOPS

3-morpholino-l-prop anesulfonate

G

guanidinophenyl

GM

(guanidinomethyl)phenyl

pNA

p-nitroanilide

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References

  1. Bieth J, Weremath CG (1970) The action of elastase onp-nitroanilide substrates. Biochem Biophys Res Commun 53: 383–390Google Scholar
  2. Brandt KG, Himoe A, Hess GP (1967) Investigations of the chymotrypsin-catalyzed hydrolysis of specific substrates. J Biol Chem 242: 3973–3982PubMedGoogle Scholar
  3. Brot FE, Bender ML (1969) Use of the specificity constant ofα-chymotrypsin. J Am Chem Soc 91: 7187–7191Google Scholar
  4. Chase T, Jr., Shaw E (1967)p-Nitrophenylp′-guanidinobenzoate-HCI: a new active site titrant for trypsin. Biochem Biophys Res Commun 29: 508–514Google Scholar
  5. Itoh K, Sekizaki H, Toyota E, Fujiwara N, Tanizawa K (1996) Application of inverse substrates to trypsin-catalyzed peptide synthesis. Bioorg Chem 24: 59–68Google Scholar
  6. Kawai M, Nagai U (1982) Upon DNP-aromatic rule. Effect of chromophore exchange on the CD spectra ofN-DNP-arylalkylamines. Bull Chem Soc Jpn 55: 1327–1328Google Scholar
  7. Nakano M, Tanizawa K, Nozawa M, Kanaoka Y (1980) Efficient tryptic hydrolysis of aryl esters with a cationic center in the leaving group. Further characterization of “inverse substrates”. Chem Pharm Bull 28: 2212–2216Google Scholar
  8. Nakatsuka T, Sasaki T, Kaiser ET (1987) Peptide segment coupling catalyzed by the semisynthetic enzyme thiolsubtilisin. J Am Chem Soc 109: 3808–3810Google Scholar
  9. Nozawa M, Tanizawa K, Kanaoka Y (1980) “Inverse substrates” for trypsin-like enzymes. J Pharmacobio-Dyn 3: 213–219PubMedGoogle Scholar
  10. Okada Y, Tsuda Y, Hirata A, Nagamatsu Y, Okamoto U (1982) Synthesis of chromogenic substrates specific for human spleen fibrinolytic protenase (SFP) and human leukocyte elastase (LE). Chem Pharm Bull 30: 4060–4068PubMedGoogle Scholar
  11. Ryan TJ, Fenton JW, II, Chang T-1, Feinman RD (1977) Specificity of thrombin: evidence for selectivity in acylation rather than binding forp-nitrophenylα-amino-p-toluate. Biochemistry 15: 1337–1341Google Scholar
  12. Schellenberger V, Jakubke H-D (1991) Protease-catalyzed kinetically controlled peptide synthesis. Angew Chem Int Ed Engl 30: 1437–1449Google Scholar
  13. Sekizaki H, Itoh K, Toyota E, Tanizawa K (1996a) Synthesis and tryptic hydrolysis ofp-guanidinophenyl esters derived from amino acids and peptides. Chem Pharm Bull 44: 1577–1579PubMedGoogle Scholar
  14. Sekizaki H, Itoh K, Toyota E, Tanizawa K (1996b) Trypsin-catalyzed peptide synthesis with variousp-guanidinophenyl esters as acyl donors. Chem Pharm Bull 44: 1585–1587PubMedGoogle Scholar
  15. Sekizaki H, Itoh K, Toyota E, Tanizawa K (1997) Enzymatic coupling of α, α-dialkyl amino acids using inverse substrates as acyl donors. Tetrahedron Lett 38: 1777–1780Google Scholar
  16. Sekizaki H, Itoh K, Toyota E, Tanizawa K (1998) Enzymatic peptide synthesis withp-guanidinophenyl andp-(guanidinomethyl)phenyl esters as acyl donors. Chem Pharm Bull 46: 846–849PubMedGoogle Scholar
  17. Tanizawa K, Kasaba Y, Kanaoka Y (1977) „Inverse substrates” for trypsin. Efficient enzymatic hydrolysis of certain esters with a cationic center in the leaving group. J Am Chem Soc 99: 4485–4488PubMedGoogle Scholar
  18. Tsuzuki H, Oka T, Morihara K (1980) Coupling between Cbz-Arg-OH and Leu-X catalyzed by trypsin and papain. J Biochem 88: 669–675PubMedGoogle Scholar
  19. Wong C-H (1989) Enzymatic catalysts in organic synthesis. Science 244: 1145–1152PubMedGoogle Scholar
  20. Yokosawa H, Hanba Y, Ishii S (1976) Affinity chromatography of trypsin and related enzymes III. Purification ofStreptomyces griseus trypsin using an affinity adsorbent containing a tryptic digest of protamine as a ligand. J Biochem 79: 757–763PubMedGoogle Scholar
  21. Yoshida N, Sasaki A, Inoue H (1971) An anionic trypsin-like enzyme fromStreptomyces eythreus. FEBS Lett 15: 129–132PubMedGoogle Scholar

Copyright information

© Springer-Verlag 1999 1999

Authors and Affiliations

  • H. Sekizaki
    • 1
  • K. Itoh
    • 1
  • E. Toyota
    • 1
  • K. Tanizawa
    • 1
  1. 1.Faculty of Pharmaceutical SciencesHealth Sciences University of HokkaidoHokkaidoJapan

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