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The Molecular Basis of Heat-Stable Enterotoxin for Vaccine Development and Cancer Cell Detection

Heat-stable enterotoxin (ST(a)) produced by Enterotoxigenic E. coli is responsible for causing acute diarrhea in infants in developing countries. However, the chemical synthesis of ST(a) peptides with the native conformation and the correct intra-molecular disulfide bonds is a major hurdle for vacci...

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Autores principales: Goto, Masaya, Yoshino, Shinya, Hiroshima, Kyona, Kawakami, Toru, Murota, Kaeko, Shimamoto, Shigeru, Hidaka, Yuji
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9920858/
https://www.ncbi.nlm.nih.gov/pubmed/36770798
http://dx.doi.org/10.3390/molecules28031128
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author Goto, Masaya
Yoshino, Shinya
Hiroshima, Kyona
Kawakami, Toru
Murota, Kaeko
Shimamoto, Shigeru
Hidaka, Yuji
author_facet Goto, Masaya
Yoshino, Shinya
Hiroshima, Kyona
Kawakami, Toru
Murota, Kaeko
Shimamoto, Shigeru
Hidaka, Yuji
author_sort Goto, Masaya
collection PubMed
description Heat-stable enterotoxin (ST(a)) produced by Enterotoxigenic E. coli is responsible for causing acute diarrhea in infants in developing countries. However, the chemical synthesis of ST(a) peptides with the native conformation and the correct intra-molecular disulfide bonds is a major hurdle for vaccine development. To address this issue, we herein report on the design and preparation of ST(a) analogs and a convenient chemical method for obtaining ST(a) molecules with the correct conformation. To develop an ST(a) vaccine, we focused on a structure in a type II β-turn in the ST(a) molecule and introduced a D-Lys residue as a conjugation site for carrier proteins. In addition, the -Glu-Leu- sequence in the ST(a) molecule was replaced with a -Asp-Val- sequence to decrease the toxic activity of the peptide to make it more amenable for use in vaccinations. To solve several issues associated with the synthesis of ST(a), such as the formation of non-native disulfide isomers, the native disulfide pairings were regioselectively formed in a stepwise manner. A native form or topological isomer of the designed ST(a) peptide, which possesses a right-handed or a left-handed spiral structure, respectively, were synthesized in high synthetic yields. The conformation of the synthetic ST(a) peptide was also confirmed by CD and NMR spectroscopy. To further utilize the designed ST(a) peptide, it was labeled with fluorescein for fluorescent detection, since recent studies have also focused on the use of ST(a) for detecting cancer cells, such as Caco-2 and T84. The labeled ST(a) peptide was able to specifically and efficiently detect 293T cells expressing the recombinant ST(a) receptor (GC-C) protein and Caco-2 cells. The findings reported here provide an outline of the molecular basis for using ST(a) for vaccine development and in the detection of cancer cells.
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spelling pubmed-99208582023-02-12 The Molecular Basis of Heat-Stable Enterotoxin for Vaccine Development and Cancer Cell Detection Goto, Masaya Yoshino, Shinya Hiroshima, Kyona Kawakami, Toru Murota, Kaeko Shimamoto, Shigeru Hidaka, Yuji Molecules Article Heat-stable enterotoxin (ST(a)) produced by Enterotoxigenic E. coli is responsible for causing acute diarrhea in infants in developing countries. However, the chemical synthesis of ST(a) peptides with the native conformation and the correct intra-molecular disulfide bonds is a major hurdle for vaccine development. To address this issue, we herein report on the design and preparation of ST(a) analogs and a convenient chemical method for obtaining ST(a) molecules with the correct conformation. To develop an ST(a) vaccine, we focused on a structure in a type II β-turn in the ST(a) molecule and introduced a D-Lys residue as a conjugation site for carrier proteins. In addition, the -Glu-Leu- sequence in the ST(a) molecule was replaced with a -Asp-Val- sequence to decrease the toxic activity of the peptide to make it more amenable for use in vaccinations. To solve several issues associated with the synthesis of ST(a), such as the formation of non-native disulfide isomers, the native disulfide pairings were regioselectively formed in a stepwise manner. A native form or topological isomer of the designed ST(a) peptide, which possesses a right-handed or a left-handed spiral structure, respectively, were synthesized in high synthetic yields. The conformation of the synthetic ST(a) peptide was also confirmed by CD and NMR spectroscopy. To further utilize the designed ST(a) peptide, it was labeled with fluorescein for fluorescent detection, since recent studies have also focused on the use of ST(a) for detecting cancer cells, such as Caco-2 and T84. The labeled ST(a) peptide was able to specifically and efficiently detect 293T cells expressing the recombinant ST(a) receptor (GC-C) protein and Caco-2 cells. The findings reported here provide an outline of the molecular basis for using ST(a) for vaccine development and in the detection of cancer cells. MDPI 2023-01-23 /pmc/articles/PMC9920858/ /pubmed/36770798 http://dx.doi.org/10.3390/molecules28031128 Text en © 2023 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Goto, Masaya
Yoshino, Shinya
Hiroshima, Kyona
Kawakami, Toru
Murota, Kaeko
Shimamoto, Shigeru
Hidaka, Yuji
The Molecular Basis of Heat-Stable Enterotoxin for Vaccine Development and Cancer Cell Detection
title The Molecular Basis of Heat-Stable Enterotoxin for Vaccine Development and Cancer Cell Detection
title_full The Molecular Basis of Heat-Stable Enterotoxin for Vaccine Development and Cancer Cell Detection
title_fullStr The Molecular Basis of Heat-Stable Enterotoxin for Vaccine Development and Cancer Cell Detection
title_full_unstemmed The Molecular Basis of Heat-Stable Enterotoxin for Vaccine Development and Cancer Cell Detection
title_short The Molecular Basis of Heat-Stable Enterotoxin for Vaccine Development and Cancer Cell Detection
title_sort molecular basis of heat-stable enterotoxin for vaccine development and cancer cell detection
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9920858/
https://www.ncbi.nlm.nih.gov/pubmed/36770798
http://dx.doi.org/10.3390/molecules28031128
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