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Topological Regulation of the Bioactive Conformation of a Disulfide-Rich Peptide, Heat-Stable Enterotoxin.

ABSTRACT: Heat-stable enterotoxin (STa) produced by enterotoxigenic E. coli causes acute diarrhea and also can be used as a specific probe for colorectal cancer cells. STa contains three intra-molecular disulfide bonds (C1-C4, C2-C5, and C3-C6 connectivity). The chemical synthesis of STa provided not only the native type of STa but also a topological isomer that had the native disulfide pairings. Interestingly, the activity of the topological isomer was approximately 1/10-1/2 that of the native STa. To further investigate the bioactive conformation of this molecule and the regulation of disulfide-coupled folding during its chemical syntheses, we examined the folding mechanism of STa that occurs during its chemical synthesis. The folding intermediate of STa with two disulfide bonds (C1-C4 and C3-C6) and two Cys(Acm) residues, the precursor peptide, was treated with iodine to produce a third disulfide bond under several conditions. The topological isomer was predominantly produced under all conditions tested, along with trace amounts of the native type of STa. In addition, NMR measurements indicated that the topological isomer has a left-handed spiral structure similar to that of the precursor peptide, while the native type of STa had a right-handed spiral structure. These results indicate that the order of the regioselective formation of disulfide bonds is important for the regulation of the final conformation of disulfide-rich peptides in chemical synthesis.

SUBMITTER: Shimamoto S

PROVIDER: S-EPMC7587965 | biostudies-literature | 2020 Oct

REPOSITORIES: biostudies-literature

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Topological Regulation of the Bioactive Conformation of a Disulfide-Rich Peptide, Heat-Stable Enterotoxin.

Shimamoto Shigeru S Fukutsuji Mayu M Osumi Toi T Goto Masaya M Toyoda Hiroshi H Hidaka Yuji Y

Molecules (Basel, Switzerland) 20201021 20

Heat-stable enterotoxin (STa) produced by enterotoxigenic E. coli causes acute diarrhea and also can be used as a specific probe for colorectal cancer cells. STa contains three intra-molecular disulfide bonds (C1-C4, C2-C5, and C3-C6 connectivity). The chemical synthesis of STa provided not only the native type of STa but also a topological isomer that had the native disulfide pairings. Interestingly, the activity of the topological isomer was a ...[more]

PMID: 33096591

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Project description:Enterotoxigenic Escherichia coli (ETEC) are a significant cause of childhood diarrhea in low-resource settings. ETEC are defined by the production of heat-stable enterotoxin (ST) and/or heat-labile enterotoxin (LT), which alter intracellular cyclic nucleotide signaling and cause the secretion of water and electrolytes into the intestinal lumen. ETEC take cues from chemicals (e.g., glycans, bile salts, and solutes) that may be liberated following enterotoxin activity to recognize entrance into the host. ETEC then alter the expression of surface adhesins called colonization factors (CFs) to attach to the intestinal epithelium, proliferate, and cause disease. Here, we used an in vivo model of oral ST intoxication to determine its impact on luminal ion concentrations via ICP-MS. We also used functional assays, including Western blots, qPCR, and toxin activity assays, to assess the impact of luminal ion flux on CF and toxin expression. Finally, we assessed ETEC strains with CFs CFA/I or CS6 in a streptomycin mouse model of ETEC colonization. ST causes rapid and significant increases in luminal chloride but significant decreases in luminal magnesium and iron. We confirmed that increased sodium chloride suppresses CFA/I production in ETEC H10407 but does not affect CS6 production in ETEC 214-4. CFA/I production in ETEC H10407 is increased when magnesium becomes limiting, although it does not affect CS6 production in ETEC 214-4. Iron restriction via deferoxamine induces CFA/I expression in ETEC H10407 but not CS6 expression in ETEC 214-4. We demonstrate that ST production is suppressed via iron restriction in H10407, 214-4, and over 50 other ETEC clinical isolates. Lastly, we demonstrate that the iron restriction of mice using oral deferoxamine pre-treatment extends the duration of ETEC H10407 (CFA/I+) fecal shedding while accelerating ETEC 214-4 (CS6+) fecal shedding. Combined, these data suggest that enterotoxins modulate luminal ion flux to influence ETEC virulence including toxin and CF production.

Dataset Information

Topological Regulation of the Bioactive Conformation of a Disulfide-Rich Peptide, Heat-Stable Enterotoxin.

Publications

Topological Regulation of the Bioactive Conformation of a Disulfide-Rich Peptide, Heat-Stable Enterotoxin.

Similar Datasets

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