Involvement of glutamic acid residue at position 7 in the formation of the intramolecular disulfide bond of Escherichia coli heat-stable enterotoxin Ip in vivo.
Escherichia coli heat-stable enterotoxin Ip (STIp) is a small peptide toxin composed of 18 amino acid residues containing three intramolecular disulfide bonds. We found previously that the bonds are formed by the catalysis of DsbA (a oxidoreductase) in periplasm [1]. To interact with DsbA, the STIp in periplasm must have a structure suitable as substrate. However, the amino acid residues contributing to the construction of this structure have not been elucidated. We mutated the codon for the glutamic acid at position 7 of STIp by oligonucleotide site-specific mutagenesis in vivo and analysed the STIp produced from the mutant gene. The intramolecular disulfide bonds were not formed in mutant STIp (Glu-7-->Ala), but were formed in mutant STIp (Glu-7-->Asp). Furthermore, we found that replacing the asparagine residue at position 11 and the proline residue at position 12 did not affect the disulfide bond formation of STIp. The results indicate that a negative charge at position 7 in the sequence of STIp is necessary for STIp to interact with DsbA in periplasm.
Yamanaka
H
H
Department of Biochemistry, Faculty of Pharmaceutical Sciences, Tokushima Bunri University, Yamashiro, Tokushima, 770-8514, Japan.
Nomura
T
T
Okamoto
K
K
D000595:Amino Acid Sequence; D001216:Asparagine / Q000502:physiology; D001427:Bacterial Toxins / Q000737:chemistry* / Q000235:genetics / Q000633:toxicity; D004220:Disulfides; D004768:Enterotoxins / Q000737:chemistry* / Q000235:genetics / Q000633:toxicity; D004926:Escherichia coli / Q000737:chemistry*; D029968:Escherichia coli Proteins; D018698:Glutamic Acid / Q000502:physiology*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011392:Proline / Q000502:physiology; D011993:Recombinant Fusion Proteins
D001427:Bacterial Toxins; D004220:Disulfides; D004768:Enterotoxins; D029968:Escherichia coli Proteins; D011993:Recombinant Fusion Proteins; C034151:heat stable toxin (E coli); D018698:Glutamic Acid; D001216:Asparagine; D011392:Proline