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Letters in Peptide Science

, Volume 4, Issue 3, pp 185–192 | Cite as

Improvement in the oxidative folding of endothelin-1 by a lys-Arg extension at the amino terminus: Implication of a salt bridge between Arg−1 and Asp8

  • Shigeru Kubo
  • Naoyoshi Chino
  • Kiichiro Nakajima
  • André Aumelas
  • Laurent Chiche
  • Shin-ichi Segawa
  • Haruhiko Tamaoki
  • Yuji Kobayashi
  • Terutoshi Kimura
  • Shumpei Sakakibara
Article

Summary

An amino-terminal extension of endothelin-l by the lys-Arg dipeptide in the prosequence (KR-ET-1) greatly increased the ratio of native-type to non-native-type disulfide isomer (96/4 versus 71/29) during the oxidative folding reaction. This improvement was completely abolished by substituting Asn for Asp at position 8 (D8N-KR-ET-1), whereas most of it was maintained with similar carboxamide analogues replaced at Glu10 or Asp18. Structure analyses by circular dichroism spectroscopy revealed that (i) in the carboxylate state, the α-helical content of the native-type isomer of KR-ET-l is higher than that of the native-type isomer of ET-1, while such a variation is not observed in the corresponding non-native-type isomer of KR-ET-l; and (ii) the enhanced α-helicity resulting from the Lys-Arg extension is largely diminished in D8N-KR-ET-l. From these results and our previous findings that the helical structure in KR-ET-l is stabilized by a particular salt bridge between the extended Arg−1 basic moiety and either the Asp8 or Glu10 acidic side chain in Et-1 [Aumelas, A. et al., Biochemistry, 34 (1995) 4546], we conclude that the formation of a specific salt bridge between the side chains of Arg−1 and Asp8 in KR-ET-1 is critical for the predominant generation of the native-type disulfide isomer, probably because it stabilizes the helical structure of parental ET-1.

Keywords

α-Helix Carboxamide substitution Circular dichroism Disulfide isomer KR-ET-1 Prosequence 

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Copyright information

© Kluwer Academic Publishers 1997

Authors and Affiliations

  • Shigeru Kubo
    • 1
  • Naoyoshi Chino
    • 1
  • Kiichiro Nakajima
    • 1
  • André Aumelas
    • 2
  • Laurent Chiche
    • 2
  • Shin-ichi Segawa
    • 3
  • Haruhiko Tamaoki
    • 4
  • Yuji Kobayashi
    • 4
  • Terutoshi Kimura
    • 1
  • Shumpei Sakakibara
    • 1
  1. 1.Protein Research FoundationPeptide Institute, Inc.OsakaJapan
  2. 2.Faculté de PharmacieCentre de Biochimie Structurale CNRS UMR C9955, INSERM U414Montpellier Cedex 1France
  3. 3.Department of Physics, School of ScienceKwansei Gakuin UniversityNishinomiyaJapan
  4. 4.Institute for Protein Research and Faculty of Pharmaceutical SciencesOsaka UniversitySuita OsakaJapan

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