Oxidative folding of human lysozyme

Effects of the loss of two bisulfide bonds and the introduction of a calcium-binding site

Yousuke Kurokawa, Nozomi Koganesawa, Yoshihiro Kobashigawa, Takumi Koshiba, Makoto Demura, Katsutoshi Nitta

Research output: Contribution to journalArticle

Abstract

Mutant human lysozymes (HLZ) lacking two disulfide bonds were constructed to study the importance of each disulfide bond on oxidative refolding. To avoid destabilization, a calcium-binding site was introduced. Five of the six species of two-disulfide mutants could be obtained with enzymatic activity. Based on the information obtained from refolding and unfolding experiments, the order of importance in oxidative refolding was found to be as follows: SS2(Cys30-Cysll6) > SS1(Cys6-Cys128) ≈ SS3(Cys65-Cys81) > SS4(Cys77-Cys95). Without SS2, these mutants refolded with low efficiency or did not refold at all. The bond SS2 is located in the interface of B-and D-helices, and a small hydrophobic cluster is formed near SS2. This cluster may play an important role in the folding process and stabilization, and SS2 may act as a stabilizer through its polypeptide linkage. The bond SS2 is the most important disulfide bond for oxidative folding of lysozymes.

Original languageEnglish
Pages (from-to)293-303
Number of pages11
JournalProtein Journal
Volume20
Issue number4
Publication statusPublished - 2001
Externally publishedYes

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Binding sites
Muramidase
Disulfides
Calcium
Enzymes
Binding Sites
Polypeptides
Stabilization
Experiments
Peptides

All Science Journal Classification (ASJC) codes

  • Analytical Chemistry
  • Bioengineering
  • Biochemistry
  • Organic Chemistry

Cite this

Kurokawa, Y., Koganesawa, N., Kobashigawa, Y., Koshiba, T., Demura, M., & Nitta, K. (2001). Oxidative folding of human lysozyme: Effects of the loss of two bisulfide bonds and the introduction of a calcium-binding site. Protein Journal, 20(4), 293-303.

Oxidative folding of human lysozyme : Effects of the loss of two bisulfide bonds and the introduction of a calcium-binding site. / Kurokawa, Yousuke; Koganesawa, Nozomi; Kobashigawa, Yoshihiro; Koshiba, Takumi; Demura, Makoto; Nitta, Katsutoshi.

In: Protein Journal, Vol. 20, No. 4, 2001, p. 293-303.

Research output: Contribution to journalArticle

Kurokawa, Y, Koganesawa, N, Kobashigawa, Y, Koshiba, T, Demura, M & Nitta, K 2001, 'Oxidative folding of human lysozyme: Effects of the loss of two bisulfide bonds and the introduction of a calcium-binding site', Protein Journal, vol. 20, no. 4, pp. 293-303.
Kurokawa Y, Koganesawa N, Kobashigawa Y, Koshiba T, Demura M, Nitta K. Oxidative folding of human lysozyme: Effects of the loss of two bisulfide bonds and the introduction of a calcium-binding site. Protein Journal. 2001;20(4):293-303.
Kurokawa, Yousuke ; Koganesawa, Nozomi ; Kobashigawa, Yoshihiro ; Koshiba, Takumi ; Demura, Makoto ; Nitta, Katsutoshi. / Oxidative folding of human lysozyme : Effects of the loss of two bisulfide bonds and the introduction of a calcium-binding site. In: Protein Journal. 2001 ; Vol. 20, No. 4. pp. 293-303.
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