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Immobilization of M3 Muscarinic Receptor to Rapidly Analyze Drug—Protein Interactions and Bioactive Components in a Natural Plant
Despite its increasing application in pursing potential ligands, the capacity of receptor affinity chromatography is greatly challenged as most current research studies lack a comprehensive characterization of the ligand–receptor interaction, particularly when simultaneously determining their bindin...
Autores principales: | , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10139132/ https://www.ncbi.nlm.nih.gov/pubmed/37108332 http://dx.doi.org/10.3390/ijms24087171 |
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author | Fan, Hushuai Huang, Xiaomin Zhang, Ziru Wang, Ting Wang, Ludan Zhang, Yajun |
author_facet | Fan, Hushuai Huang, Xiaomin Zhang, Ziru Wang, Ting Wang, Ludan Zhang, Yajun |
author_sort | Fan, Hushuai |
collection | PubMed |
description | Despite its increasing application in pursing potential ligands, the capacity of receptor affinity chromatography is greatly challenged as most current research studies lack a comprehensive characterization of the ligand–receptor interaction, particularly when simultaneously determining their binding thermodynamics and kinetics. This work developed an immobilized M3 muscarinic receptor (M3R) affinity column by fixing M3R on amino polystyrene microspheres via the interaction of a 6-chlorohexanoic acid linker with haloalkane dehalogenase. The efficiency of the immobilized M3R was tested by characterizing the binding thermodynamics and kinetics of three known drugs to immobilized M3R using a frontal analysis and the peak profiling method, as well as by analyzing the bioactive compounds in Daturae Flos (DF) extract. The data showed that the immobilized M3R demonstrated good specificity, stability, and competence for analyzing drug–protein interactions. The association constants of (−)-scopolamine hydrochloride, atropine sulfate, and pilocarpine to M3R were determined to be (2.39 ± 0.03) × 10(4), (3.71 ± 0.03) × 10(4), and (2.73 ± 0.04) × 10(4) M(−1), respectively, with dissociation rate constants of 27.47 ± 0.65, 14.28 ± 0.17, and 10.70 ± 0.35 min(−1), respectively. Hyoscyamine and scopolamine were verified as the bioactive compounds that bind to M3R in the DF extract. Our results suggest that the immobilized M3R method was capable of determining drug–protein binding parameters and probing specific ligands in a natural plant, thus enhancing the effectiveness of receptor affinity chromatography in diverse stages of drug discovery. |
format | Online Article Text |
id | pubmed-10139132 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-101391322023-04-28 Immobilization of M3 Muscarinic Receptor to Rapidly Analyze Drug—Protein Interactions and Bioactive Components in a Natural Plant Fan, Hushuai Huang, Xiaomin Zhang, Ziru Wang, Ting Wang, Ludan Zhang, Yajun Int J Mol Sci Article Despite its increasing application in pursing potential ligands, the capacity of receptor affinity chromatography is greatly challenged as most current research studies lack a comprehensive characterization of the ligand–receptor interaction, particularly when simultaneously determining their binding thermodynamics and kinetics. This work developed an immobilized M3 muscarinic receptor (M3R) affinity column by fixing M3R on amino polystyrene microspheres via the interaction of a 6-chlorohexanoic acid linker with haloalkane dehalogenase. The efficiency of the immobilized M3R was tested by characterizing the binding thermodynamics and kinetics of three known drugs to immobilized M3R using a frontal analysis and the peak profiling method, as well as by analyzing the bioactive compounds in Daturae Flos (DF) extract. The data showed that the immobilized M3R demonstrated good specificity, stability, and competence for analyzing drug–protein interactions. The association constants of (−)-scopolamine hydrochloride, atropine sulfate, and pilocarpine to M3R were determined to be (2.39 ± 0.03) × 10(4), (3.71 ± 0.03) × 10(4), and (2.73 ± 0.04) × 10(4) M(−1), respectively, with dissociation rate constants of 27.47 ± 0.65, 14.28 ± 0.17, and 10.70 ± 0.35 min(−1), respectively. Hyoscyamine and scopolamine were verified as the bioactive compounds that bind to M3R in the DF extract. Our results suggest that the immobilized M3R method was capable of determining drug–protein binding parameters and probing specific ligands in a natural plant, thus enhancing the effectiveness of receptor affinity chromatography in diverse stages of drug discovery. MDPI 2023-04-12 /pmc/articles/PMC10139132/ /pubmed/37108332 http://dx.doi.org/10.3390/ijms24087171 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 Fan, Hushuai Huang, Xiaomin Zhang, Ziru Wang, Ting Wang, Ludan Zhang, Yajun Immobilization of M3 Muscarinic Receptor to Rapidly Analyze Drug—Protein Interactions and Bioactive Components in a Natural Plant |
title | Immobilization of M3 Muscarinic Receptor to Rapidly Analyze Drug—Protein Interactions and Bioactive Components in a Natural Plant |
title_full | Immobilization of M3 Muscarinic Receptor to Rapidly Analyze Drug—Protein Interactions and Bioactive Components in a Natural Plant |
title_fullStr | Immobilization of M3 Muscarinic Receptor to Rapidly Analyze Drug—Protein Interactions and Bioactive Components in a Natural Plant |
title_full_unstemmed | Immobilization of M3 Muscarinic Receptor to Rapidly Analyze Drug—Protein Interactions and Bioactive Components in a Natural Plant |
title_short | Immobilization of M3 Muscarinic Receptor to Rapidly Analyze Drug—Protein Interactions and Bioactive Components in a Natural Plant |
title_sort | immobilization of m3 muscarinic receptor to rapidly analyze drug—protein interactions and bioactive components in a natural plant |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10139132/ https://www.ncbi.nlm.nih.gov/pubmed/37108332 http://dx.doi.org/10.3390/ijms24087171 |
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