Disulfide bond formation and secretion of Escherichia coli heat-stable enterotoxin II

Keinosuke Okamoto, T. Baba, H. Yamanaka, N. Akashi, Y. Fujii

Research output: Contribution to journalArticle

44 Citations (Scopus)

Abstract

The Escherichia coli heat-stable enterotoxin II (STII) is a typical extracellular toxin consisting of 48 amino acid residues, of which 4 are cysteine. There are two disulfide bonds, one between Cys-10 and Cys-48 and one between Cys-21 and Cys-36. We examined the involvement of Dsba in the formation of the disulfide bonds of STII and the role of each in the secretion of STII. A dsbA mutant was transformed with a plasmid harboring the STII gene, and STII was not detected either in the cells or in the culture supernatant. Reducing the level of STII brought about the dsbA mutation restored by introducing the wild-type dsbA gene into the mutant strain. These results showed that DsbA is involved in forming the disulfide bonds of STII and that STII without these disulfide bonds is degraded during secretion. We substituted these four cysteine residues in vivo by oligonucleotide-directed site-specific mutagenesis. The amino acid sequence of the purified STII(C48S) and pulse-chase studies revealed that two intermolecular disulfide bonds must be formed to be efficiently secreted and that cleavage between amino acid residues 14 and 15 is probably the first step in the proteolytic degradation of STII.

Original languageEnglish
Pages (from-to)4579-4586
Number of pages8
JournalJournal of Bacteriology
Volume177
Issue number16
Publication statusPublished - 1995
Externally publishedYes

Fingerprint

Enterotoxins
Disulfides
Escherichia coli
Cysteine
heat stable toxin (E coli)
Amino Acids
Site-Directed Mutagenesis
Oligonucleotides
Genes
Amino Acid Sequence
Plasmids
Mutation

ASJC Scopus subject areas

  • Applied Microbiology and Biotechnology
  • Immunology

Cite this

Okamoto, K., Baba, T., Yamanaka, H., Akashi, N., & Fujii, Y. (1995). Disulfide bond formation and secretion of Escherichia coli heat-stable enterotoxin II. Journal of Bacteriology, 177(16), 4579-4586.

Disulfide bond formation and secretion of Escherichia coli heat-stable enterotoxin II. / Okamoto, Keinosuke; Baba, T.; Yamanaka, H.; Akashi, N.; Fujii, Y.

In: Journal of Bacteriology, Vol. 177, No. 16, 1995, p. 4579-4586.

Research output: Contribution to journalArticle

Okamoto, K, Baba, T, Yamanaka, H, Akashi, N & Fujii, Y 1995, 'Disulfide bond formation and secretion of Escherichia coli heat-stable enterotoxin II', Journal of Bacteriology, vol. 177, no. 16, pp. 4579-4586.
Okamoto, Keinosuke ; Baba, T. ; Yamanaka, H. ; Akashi, N. ; Fujii, Y. / Disulfide bond formation and secretion of Escherichia coli heat-stable enterotoxin II. In: Journal of Bacteriology. 1995 ; Vol. 177, No. 16. pp. 4579-4586.
@article{6b8a7293ec1a4ee6800ea12872725161,
title = "Disulfide bond formation and secretion of Escherichia coli heat-stable enterotoxin II",
abstract = "The Escherichia coli heat-stable enterotoxin II (STII) is a typical extracellular toxin consisting of 48 amino acid residues, of which 4 are cysteine. There are two disulfide bonds, one between Cys-10 and Cys-48 and one between Cys-21 and Cys-36. We examined the involvement of Dsba in the formation of the disulfide bonds of STII and the role of each in the secretion of STII. A dsbA mutant was transformed with a plasmid harboring the STII gene, and STII was not detected either in the cells or in the culture supernatant. Reducing the level of STII brought about the dsbA mutation restored by introducing the wild-type dsbA gene into the mutant strain. These results showed that DsbA is involved in forming the disulfide bonds of STII and that STII without these disulfide bonds is degraded during secretion. We substituted these four cysteine residues in vivo by oligonucleotide-directed site-specific mutagenesis. The amino acid sequence of the purified STII(C48S) and pulse-chase studies revealed that two intermolecular disulfide bonds must be formed to be efficiently secreted and that cleavage between amino acid residues 14 and 15 is probably the first step in the proteolytic degradation of STII.",
author = "Keinosuke Okamoto and T. Baba and H. Yamanaka and N. Akashi and Y. Fujii",
year = "1995",
language = "English",
volume = "177",
pages = "4579--4586",
journal = "Journal of Bacteriology",
issn = "0021-9193",
publisher = "American Society for Microbiology",
number = "16",

}

TY - JOUR

T1 - Disulfide bond formation and secretion of Escherichia coli heat-stable enterotoxin II

AU - Okamoto, Keinosuke

AU - Baba, T.

AU - Yamanaka, H.

AU - Akashi, N.

AU - Fujii, Y.

PY - 1995

Y1 - 1995

N2 - The Escherichia coli heat-stable enterotoxin II (STII) is a typical extracellular toxin consisting of 48 amino acid residues, of which 4 are cysteine. There are two disulfide bonds, one between Cys-10 and Cys-48 and one between Cys-21 and Cys-36. We examined the involvement of Dsba in the formation of the disulfide bonds of STII and the role of each in the secretion of STII. A dsbA mutant was transformed with a plasmid harboring the STII gene, and STII was not detected either in the cells or in the culture supernatant. Reducing the level of STII brought about the dsbA mutation restored by introducing the wild-type dsbA gene into the mutant strain. These results showed that DsbA is involved in forming the disulfide bonds of STII and that STII without these disulfide bonds is degraded during secretion. We substituted these four cysteine residues in vivo by oligonucleotide-directed site-specific mutagenesis. The amino acid sequence of the purified STII(C48S) and pulse-chase studies revealed that two intermolecular disulfide bonds must be formed to be efficiently secreted and that cleavage between amino acid residues 14 and 15 is probably the first step in the proteolytic degradation of STII.

AB - The Escherichia coli heat-stable enterotoxin II (STII) is a typical extracellular toxin consisting of 48 amino acid residues, of which 4 are cysteine. There are two disulfide bonds, one between Cys-10 and Cys-48 and one between Cys-21 and Cys-36. We examined the involvement of Dsba in the formation of the disulfide bonds of STII and the role of each in the secretion of STII. A dsbA mutant was transformed with a plasmid harboring the STII gene, and STII was not detected either in the cells or in the culture supernatant. Reducing the level of STII brought about the dsbA mutation restored by introducing the wild-type dsbA gene into the mutant strain. These results showed that DsbA is involved in forming the disulfide bonds of STII and that STII without these disulfide bonds is degraded during secretion. We substituted these four cysteine residues in vivo by oligonucleotide-directed site-specific mutagenesis. The amino acid sequence of the purified STII(C48S) and pulse-chase studies revealed that two intermolecular disulfide bonds must be formed to be efficiently secreted and that cleavage between amino acid residues 14 and 15 is probably the first step in the proteolytic degradation of STII.

UR - http://www.scopus.com/inward/record.url?scp=0029118246&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0029118246&partnerID=8YFLogxK

M3 - Article

VL - 177

SP - 4579

EP - 4586

JO - Journal of Bacteriology

JF - Journal of Bacteriology

SN - 0021-9193

IS - 16

ER -