Cargando…

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....

Descripción completa

Detalles Bibliográficos
Autores principales: Yamanaka, Daisuke, Kurita, Suzuka, Hanayama, Yuka, Adachi, Yoshiyuki
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2021
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
_version_ 1783657307958149120
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
work_keys_str_mv AT yamanakadaisuke splitenzymebasedbiosensorsforstructuralcharacterizationofsolubleandinsolublebglucans
AT kuritasuzuka splitenzymebasedbiosensorsforstructuralcharacterizationofsolubleandinsolublebglucans
AT hanayamayuka splitenzymebasedbiosensorsforstructuralcharacterizationofsolubleandinsolublebglucans
AT adachiyoshiyuki splitenzymebasedbiosensorsforstructuralcharacterizationofsolubleandinsolublebglucans