Screening method toward ClbP-specific inhibitors
Abstract Background Colibactin is a genotoxin produced by Escherichia coli and other Enterobacteriaceae that is believed to increase the risk of colorectal cancer (CRC) of their symbiosis hosts, including human. A peptidase ClbP is the key enzyme for activation of colibactin. Inhibition of ClbP is c...
| Main Authors: | , , , , , , , , |
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| Format: | Article |
| Language: | English |
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BMC
2023-02-01
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| Series: | Genes and Environment |
| Subjects: | |
| Online Access: | https://doi.org/10.1186/s41021-023-00264-7 |
| _version_ | 1797863540877426688 |
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| author | Tao Zhou Takayuki Ando Akihiro Kudo Michio Sato Noriyuki Miyoshi Michihiro Mutoh Hideki Ishikawa Keiji Wakabayashi Kenji Watanabe |
| author_facet | Tao Zhou Takayuki Ando Akihiro Kudo Michio Sato Noriyuki Miyoshi Michihiro Mutoh Hideki Ishikawa Keiji Wakabayashi Kenji Watanabe |
| author_sort | Tao Zhou |
| collection | DOAJ |
| description | Abstract Background Colibactin is a genotoxin produced by Escherichia coli and other Enterobacteriaceae that is believed to increase the risk of colorectal cancer (CRC) of their symbiosis hosts, including human. A peptidase ClbP is the key enzyme for activation of colibactin. Inhibition of ClbP is considered to impede maturation of precolibactin into genotoxic colibactin. Therefore, ClbP-specific inhibitors could potentially prevent the onset of CRC, one of the leading causes of cancer-related deaths in the world. This study intends to establish an efficient screening system for identifying inhibitors that are specific to ClbP. Methods Two types of assays were applied in the screening procedure: a probe assay and an LC–MS assay. For the probe assay, we employed the synthesized probe which we described in our previous report. This probe can be hydrolyzed efficiently by ClbP to release a fluorophore. Hence it was applied here for detection of inhibition of ClbP. For the LC–MS assay, formation of the byproduct of precolibactin maturation process, N-myristoyl-D-asparagine, was quantified using a liquid chromatography–mass spectrometry (LC–MS) technique. The probe assay can be performed much faster, while the LC–MS assay is more accurate. Therefore, our method employed the two assays in sequence to screen a large number of compounds for inhibition of ClbP. Results A library of 67,965 standard compounds was evaluated by the screening method established in the current study, and one compound was found to show a moderate inhibitory activity against ClbP. Conclusion A simple screening method for ClbP-specific inhibitors was established. It was proven to be reliable and is believed to be useful in developing potential prophylactic agents for CRC. |
| first_indexed | 2024-04-09T22:37:08Z |
| format | Article |
| id | doaj.art-704efcc564b04bcf83a51dce81db4a28 |
| institution | Directory Open Access Journal |
| issn | 1880-7062 |
| language | English |
| last_indexed | 2024-04-09T22:37:08Z |
| publishDate | 2023-02-01 |
| publisher | BMC |
| record_format | Article |
| series | Genes and Environment |
| spelling | doaj.art-704efcc564b04bcf83a51dce81db4a282023-03-22T12:23:37ZengBMCGenes and Environment1880-70622023-02-014511710.1186/s41021-023-00264-7Screening method toward ClbP-specific inhibitorsTao Zhou0Takayuki Ando1Akihiro Kudo2Michio Sato3Noriyuki Miyoshi4Michihiro Mutoh5Hideki Ishikawa6Keiji Wakabayashi7Kenji Watanabe8Adenoprevent Co., Ltd.Department of Pharmaceutical and Food Science, Shizuoka Institution of Environment and HygieneDepartment of Pharmaceutical and Food Science, Shizuoka Institution of Environment and HygieneDepartment of Pharmaceutical Sciences, University of ShizuokaGraduate School of Integrated Pharmaceutical and Nutritional Sciences, University of ShizuokaDepartment of Molecular-Targeting Cancer Prevention, Kyoto Prefectural University of MedicineDepartment of Molecular-Targeting Cancer Prevention, Kyoto Prefectural University of MedicineGraduate School of Nutritional and Environmental Sciences, University of ShizuokaAdenoprevent Co., Ltd.Abstract Background Colibactin is a genotoxin produced by Escherichia coli and other Enterobacteriaceae that is believed to increase the risk of colorectal cancer (CRC) of their symbiosis hosts, including human. A peptidase ClbP is the key enzyme for activation of colibactin. Inhibition of ClbP is considered to impede maturation of precolibactin into genotoxic colibactin. Therefore, ClbP-specific inhibitors could potentially prevent the onset of CRC, one of the leading causes of cancer-related deaths in the world. This study intends to establish an efficient screening system for identifying inhibitors that are specific to ClbP. Methods Two types of assays were applied in the screening procedure: a probe assay and an LC–MS assay. For the probe assay, we employed the synthesized probe which we described in our previous report. This probe can be hydrolyzed efficiently by ClbP to release a fluorophore. Hence it was applied here for detection of inhibition of ClbP. For the LC–MS assay, formation of the byproduct of precolibactin maturation process, N-myristoyl-D-asparagine, was quantified using a liquid chromatography–mass spectrometry (LC–MS) technique. The probe assay can be performed much faster, while the LC–MS assay is more accurate. Therefore, our method employed the two assays in sequence to screen a large number of compounds for inhibition of ClbP. Results A library of 67,965 standard compounds was evaluated by the screening method established in the current study, and one compound was found to show a moderate inhibitory activity against ClbP. Conclusion A simple screening method for ClbP-specific inhibitors was established. It was proven to be reliable and is believed to be useful in developing potential prophylactic agents for CRC.https://doi.org/10.1186/s41021-023-00264-7ColibactinClbPInhibitorScreeningFluorescentProbe |
| spellingShingle | Tao Zhou Takayuki Ando Akihiro Kudo Michio Sato Noriyuki Miyoshi Michihiro Mutoh Hideki Ishikawa Keiji Wakabayashi Kenji Watanabe Screening method toward ClbP-specific inhibitors Genes and Environment Colibactin ClbP Inhibitor Screening Fluorescent Probe |
| title | Screening method toward ClbP-specific inhibitors |
| title_full | Screening method toward ClbP-specific inhibitors |
| title_fullStr | Screening method toward ClbP-specific inhibitors |
| title_full_unstemmed | Screening method toward ClbP-specific inhibitors |
| title_short | Screening method toward ClbP-specific inhibitors |
| title_sort | screening method toward clbp specific inhibitors |
| topic | Colibactin ClbP Inhibitor Screening Fluorescent Probe |
| url | https://doi.org/10.1186/s41021-023-00264-7 |
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