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Switch-on Fluorescence Analysis of Protease Activity with the Assistance of a Nickel Ion-Nitrilotriacetic Acid-Conjugated Magnetic Nanoparticle
Heterogeneous protease biosensors show high sensitivity and selectivity but usually require the immobilization of peptide substrates on a solid interface. Such methods exhibit the disadvantages of complex immobilization steps and low enzymatic efficiency induced by steric hindrance. In this work, we...
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/PMC10144723/ https://www.ncbi.nlm.nih.gov/pubmed/37110659 http://dx.doi.org/10.3390/molecules28083426 |
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author | Ma, Xiaohua Lv, Yingxin Liu, Panpan Hao, Yuanqiang Xia, Ning |
author_facet | Ma, Xiaohua Lv, Yingxin Liu, Panpan Hao, Yuanqiang Xia, Ning |
author_sort | Ma, Xiaohua |
collection | PubMed |
description | Heterogeneous protease biosensors show high sensitivity and selectivity but usually require the immobilization of peptide substrates on a solid interface. Such methods exhibit the disadvantages of complex immobilization steps and low enzymatic efficiency induced by steric hindrance. In this work, we proposed an immobilization-free strategy for protease detection with high simplicity, sensitivity and selectivity. Specifically, a single-labeled peptide with oligohistidine-tag (His-tag) was designed as the protease substrate, which can be captured by a nickel ion-nitrilotriacetic acid (Ni-NTA)-conjugated magnetic nanoparticle (MNP) through the coordination interaction between His-tag and Ni-NTA. When the peptide was digested by protease in a homogeneous solution, the signal-labeled segment was released from the substrate. The unreacted peptide substrates could be removed by Ni-NTA-MNP, and the released segments remained in solution to emit strong fluorescence. The method was used to determine protease of caspase-3 with a low detection limit (4 pg/mL). By changing the peptide sequence and signal reporters, the proposal could be used to develop novel homogeneous biosensors for the detection of other proteases. |
format | Online Article Text |
id | pubmed-10144723 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-101447232023-04-29 Switch-on Fluorescence Analysis of Protease Activity with the Assistance of a Nickel Ion-Nitrilotriacetic Acid-Conjugated Magnetic Nanoparticle Ma, Xiaohua Lv, Yingxin Liu, Panpan Hao, Yuanqiang Xia, Ning Molecules Communication Heterogeneous protease biosensors show high sensitivity and selectivity but usually require the immobilization of peptide substrates on a solid interface. Such methods exhibit the disadvantages of complex immobilization steps and low enzymatic efficiency induced by steric hindrance. In this work, we proposed an immobilization-free strategy for protease detection with high simplicity, sensitivity and selectivity. Specifically, a single-labeled peptide with oligohistidine-tag (His-tag) was designed as the protease substrate, which can be captured by a nickel ion-nitrilotriacetic acid (Ni-NTA)-conjugated magnetic nanoparticle (MNP) through the coordination interaction between His-tag and Ni-NTA. When the peptide was digested by protease in a homogeneous solution, the signal-labeled segment was released from the substrate. The unreacted peptide substrates could be removed by Ni-NTA-MNP, and the released segments remained in solution to emit strong fluorescence. The method was used to determine protease of caspase-3 with a low detection limit (4 pg/mL). By changing the peptide sequence and signal reporters, the proposal could be used to develop novel homogeneous biosensors for the detection of other proteases. MDPI 2023-04-13 /pmc/articles/PMC10144723/ /pubmed/37110659 http://dx.doi.org/10.3390/molecules28083426 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 | Communication Ma, Xiaohua Lv, Yingxin Liu, Panpan Hao, Yuanqiang Xia, Ning Switch-on Fluorescence Analysis of Protease Activity with the Assistance of a Nickel Ion-Nitrilotriacetic Acid-Conjugated Magnetic Nanoparticle |
title | Switch-on Fluorescence Analysis of Protease Activity with the Assistance of a Nickel Ion-Nitrilotriacetic Acid-Conjugated Magnetic Nanoparticle |
title_full | Switch-on Fluorescence Analysis of Protease Activity with the Assistance of a Nickel Ion-Nitrilotriacetic Acid-Conjugated Magnetic Nanoparticle |
title_fullStr | Switch-on Fluorescence Analysis of Protease Activity with the Assistance of a Nickel Ion-Nitrilotriacetic Acid-Conjugated Magnetic Nanoparticle |
title_full_unstemmed | Switch-on Fluorescence Analysis of Protease Activity with the Assistance of a Nickel Ion-Nitrilotriacetic Acid-Conjugated Magnetic Nanoparticle |
title_short | Switch-on Fluorescence Analysis of Protease Activity with the Assistance of a Nickel Ion-Nitrilotriacetic Acid-Conjugated Magnetic Nanoparticle |
title_sort | switch-on fluorescence analysis of protease activity with the assistance of a nickel ion-nitrilotriacetic acid-conjugated magnetic nanoparticle |
topic | Communication |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10144723/ https://www.ncbi.nlm.nih.gov/pubmed/37110659 http://dx.doi.org/10.3390/molecules28083426 |
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