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Split Enzyme-Based Biosensors for Structural Characterization of Soluble and Insoluble β-Glucans
β-Glucan is widely distributed in various plants and microorganisms and is composed of β-1,3-linked d-glucose units. It may have a branched short or long side chain of glucose units with β-1,6- or β-1,4-linkage. Numerous studies have investigated different β-glucans and revealed their bioactivities....
Autores principales: | , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2021
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7915705/ https://www.ncbi.nlm.nih.gov/pubmed/33557290 http://dx.doi.org/10.3390/ijms22041576 |
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author | Yamanaka, Daisuke Kurita, Suzuka Hanayama, Yuka Adachi, Yoshiyuki |
author_facet | Yamanaka, Daisuke Kurita, Suzuka Hanayama, Yuka Adachi, Yoshiyuki |
author_sort | Yamanaka, Daisuke |
collection | PubMed |
description | β-Glucan is widely distributed in various plants and microorganisms and is composed of β-1,3-linked d-glucose units. It may have a branched short or long side chain of glucose units with β-1,6- or β-1,4-linkage. Numerous studies have investigated different β-glucans and revealed their bioactivities. To understand the structure-function relationship of β-glucan, we constructed a split-luciferase complementation assay for the structural analysis of long-chain β-1,6-branched β-1,3-glucan. The N- and C-terminal fragments of luciferase from deep-sea shrimp were fused to insect-derived β-1,3-glucan recognition protein and fungal endo-β-1,6-glucanase (Neg1)-derived β-1,6-glucan recognition protein, respectively. In this approach, two β-glucan recognition proteins bound to β-glucan molecules come into close proximity, resulting in the assembly of the full-length reporter enzyme and induction of transient luciferase activity, indicative of the structure of β-glucan. To test the applicability of this assay, β-glucan and two β-glucan recognition proteins were mixed, resulting in an increase in the luminescence intensity in a β-1,3-glucan with a long polymer of β-1,6-glucan in a dose-dependent manner. This simple test also allows the monitoring of real-time changes in the side chain structure and serves as a convenient method to distinguish between β-1,3-glucan and long-chain β-1,6-branched β-1,3-glucan in various soluble and insoluble β-glucans. |
format | Online Article Text |
id | pubmed-7915705 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-79157052021-03-01 Split Enzyme-Based Biosensors for Structural Characterization of Soluble and Insoluble β-Glucans Yamanaka, Daisuke Kurita, Suzuka Hanayama, Yuka Adachi, Yoshiyuki Int J Mol Sci Article β-Glucan is widely distributed in various plants and microorganisms and is composed of β-1,3-linked d-glucose units. It may have a branched short or long side chain of glucose units with β-1,6- or β-1,4-linkage. Numerous studies have investigated different β-glucans and revealed their bioactivities. To understand the structure-function relationship of β-glucan, we constructed a split-luciferase complementation assay for the structural analysis of long-chain β-1,6-branched β-1,3-glucan. The N- and C-terminal fragments of luciferase from deep-sea shrimp were fused to insect-derived β-1,3-glucan recognition protein and fungal endo-β-1,6-glucanase (Neg1)-derived β-1,6-glucan recognition protein, respectively. In this approach, two β-glucan recognition proteins bound to β-glucan molecules come into close proximity, resulting in the assembly of the full-length reporter enzyme and induction of transient luciferase activity, indicative of the structure of β-glucan. To test the applicability of this assay, β-glucan and two β-glucan recognition proteins were mixed, resulting in an increase in the luminescence intensity in a β-1,3-glucan with a long polymer of β-1,6-glucan in a dose-dependent manner. This simple test also allows the monitoring of real-time changes in the side chain structure and serves as a convenient method to distinguish between β-1,3-glucan and long-chain β-1,6-branched β-1,3-glucan in various soluble and insoluble β-glucans. MDPI 2021-02-04 /pmc/articles/PMC7915705/ /pubmed/33557290 http://dx.doi.org/10.3390/ijms22041576 Text en © 2021 by the authors. 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 (http://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Yamanaka, Daisuke Kurita, Suzuka Hanayama, Yuka Adachi, Yoshiyuki Split Enzyme-Based Biosensors for Structural Characterization of Soluble and Insoluble β-Glucans |
title | Split Enzyme-Based Biosensors for Structural Characterization of Soluble and Insoluble β-Glucans |
title_full | Split Enzyme-Based Biosensors for Structural Characterization of Soluble and Insoluble β-Glucans |
title_fullStr | Split Enzyme-Based Biosensors for Structural Characterization of Soluble and Insoluble β-Glucans |
title_full_unstemmed | Split Enzyme-Based Biosensors for Structural Characterization of Soluble and Insoluble β-Glucans |
title_short | Split Enzyme-Based Biosensors for Structural Characterization of Soluble and Insoluble β-Glucans |
title_sort | split enzyme-based biosensors for structural characterization of soluble and insoluble β-glucans |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7915705/ https://www.ncbi.nlm.nih.gov/pubmed/33557290 http://dx.doi.org/10.3390/ijms22041576 |
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