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A label-free LC/MS-based enzymatic activity assay for the detection of PDE5A inhibitors

Phosphodiesterase type 5 (PDE5), a cyclic nucleotide phosphodiesterase, controls the duration of the cyclic guanosine monophosphate (cGMP) signal by hydrolyzing cGMP to GMP. Inhibiting the activity of PDE5A has proven to be an effective strategy for treating pulmonary arterial hypertension and erect...

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Autores principales: Ma, Yufeng, Zhang, Fengsen, Zhong, Yijing, Huang, Yongchun, Yixizhuoma, Jia, Qiangqiang, Zhang, Shoude
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9968969/
https://www.ncbi.nlm.nih.gov/pubmed/36860644
http://dx.doi.org/10.3389/fchem.2023.1097027
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author Ma, Yufeng
Zhang, Fengsen
Zhong, Yijing
Huang, Yongchun
Yixizhuoma,
Jia, Qiangqiang
Zhang, Shoude
author_facet Ma, Yufeng
Zhang, Fengsen
Zhong, Yijing
Huang, Yongchun
Yixizhuoma,
Jia, Qiangqiang
Zhang, Shoude
author_sort Ma, Yufeng
collection PubMed
description Phosphodiesterase type 5 (PDE5), a cyclic nucleotide phosphodiesterase, controls the duration of the cyclic guanosine monophosphate (cGMP) signal by hydrolyzing cGMP to GMP. Inhibiting the activity of PDE5A has proven to be an effective strategy for treating pulmonary arterial hypertension and erectile dysfunction. Current enzymatic activity assay methods for PDE5A mainly use fluorescent or isotope-labeled substrates, which are expensive and inconvenient. Here, we developed an LC/MS-based enzymatic activity assay for PDE5A without labeling, which detects the enzymatic activity of PDE5A by quantifying the substrate cGMP and product GMP at a concentration of 100 nM. The accuracy of this method was verified by a fluorescently labeled substrate. Moreover, a new inhibitor of PDE5A was identified by this method and virtual screening. It inhibited PDE5A with an IC(50) value of 870 nM. Overall, the proposed strategy provides a new method for screening PDE5A inhibitors.
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spelling pubmed-99689692023-02-28 A label-free LC/MS-based enzymatic activity assay for the detection of PDE5A inhibitors Ma, Yufeng Zhang, Fengsen Zhong, Yijing Huang, Yongchun Yixizhuoma, Jia, Qiangqiang Zhang, Shoude Front Chem Chemistry Phosphodiesterase type 5 (PDE5), a cyclic nucleotide phosphodiesterase, controls the duration of the cyclic guanosine monophosphate (cGMP) signal by hydrolyzing cGMP to GMP. Inhibiting the activity of PDE5A has proven to be an effective strategy for treating pulmonary arterial hypertension and erectile dysfunction. Current enzymatic activity assay methods for PDE5A mainly use fluorescent or isotope-labeled substrates, which are expensive and inconvenient. Here, we developed an LC/MS-based enzymatic activity assay for PDE5A without labeling, which detects the enzymatic activity of PDE5A by quantifying the substrate cGMP and product GMP at a concentration of 100 nM. The accuracy of this method was verified by a fluorescently labeled substrate. Moreover, a new inhibitor of PDE5A was identified by this method and virtual screening. It inhibited PDE5A with an IC(50) value of 870 nM. Overall, the proposed strategy provides a new method for screening PDE5A inhibitors. Frontiers Media S.A. 2023-02-13 /pmc/articles/PMC9968969/ /pubmed/36860644 http://dx.doi.org/10.3389/fchem.2023.1097027 Text en Copyright © 2023 Ma, Zhang, Zhong, Huang, Yixizhuoma, Jia and Zhang. https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Chemistry
Ma, Yufeng
Zhang, Fengsen
Zhong, Yijing
Huang, Yongchun
Yixizhuoma,
Jia, Qiangqiang
Zhang, Shoude
A label-free LC/MS-based enzymatic activity assay for the detection of PDE5A inhibitors
title A label-free LC/MS-based enzymatic activity assay for the detection of PDE5A inhibitors
title_full A label-free LC/MS-based enzymatic activity assay for the detection of PDE5A inhibitors
title_fullStr A label-free LC/MS-based enzymatic activity assay for the detection of PDE5A inhibitors
title_full_unstemmed A label-free LC/MS-based enzymatic activity assay for the detection of PDE5A inhibitors
title_short A label-free LC/MS-based enzymatic activity assay for the detection of PDE5A inhibitors
title_sort label-free lc/ms-based enzymatic activity assay for the detection of pde5a inhibitors
topic Chemistry
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9968969/
https://www.ncbi.nlm.nih.gov/pubmed/36860644
http://dx.doi.org/10.3389/fchem.2023.1097027
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