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Functional Expression and Characterization of the Recombinant N-Acetyl-Glucosamine/N-Acetyl-Galactosamine-Specific Marine Algal Lectin BPL3
Lectins, characterized by their carbohydrate-binding ability, have extensive practical applications. However, their industrial use is limited due to impurity. Thus, quality-controlled production of recombinant lectin is necessary. In this study, the algal lectin BPL3 (Bryopsis plumosa lectin 3) was...
Autores principales: | , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
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MDPI
2018
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5793061/ https://www.ncbi.nlm.nih.gov/pubmed/29303968 http://dx.doi.org/10.3390/md16010013 |
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author | Hwang, Hyun-Ju Han, Jin-Woo Kim, Gwang Hoon Han, Jong Won |
author_facet | Hwang, Hyun-Ju Han, Jin-Woo Kim, Gwang Hoon Han, Jong Won |
author_sort | Hwang, Hyun-Ju |
collection | PubMed |
description | Lectins, characterized by their carbohydrate-binding ability, have extensive practical applications. However, their industrial use is limited due to impurity. Thus, quality-controlled production of recombinant lectin is necessary. In this study, the algal lectin BPL3 (Bryopsis plumosa lectin 3) was successfully produced using a bacterial expression system, BL21(DE3), with an artificial repeated structure (dimeric construct). Recombinant dimeric BPL3 (rD2BPL3) was confirmed by LC-MS/MS spectrometry. Expression efficiency was greater for the construct with the repeat structure (rD2BPL3) than the monomeric form (rD1BPL3). Optimal conditions for expression were 1 mM IPTG at 20 °C. Recombinant lectin was purified under denaturing conditions and refolded by the flash dilution method. Recombinant BPL3 was solubilized in 1× PBS containing 2 M urea. rD2BPL3 showed strong hemagglutination activity using human erythrocyte. rD2BPL3 had a similar sugar specificity to that of the native protein, i.e., to N-acetyl-glucosamine (GlcNAc) and N-acetyl-galactosamine (GalNAc). Glycan array results showed that recombinant BPL3 and native BPL3 exhibited different binding properties. Both showed weak binding activity to α-Man-Sp. Native BPL3 showed strong binding specificity to the alpha conformation of amino sugars, and rD2BPL3 had binding activity to the beta conformation. The process developed in this study was suitable for the quality-controlled large-scale production of recombinant lectins. |
format | Online Article Text |
id | pubmed-5793061 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2018 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-57930612018-02-06 Functional Expression and Characterization of the Recombinant N-Acetyl-Glucosamine/N-Acetyl-Galactosamine-Specific Marine Algal Lectin BPL3 Hwang, Hyun-Ju Han, Jin-Woo Kim, Gwang Hoon Han, Jong Won Mar Drugs Article Lectins, characterized by their carbohydrate-binding ability, have extensive practical applications. However, their industrial use is limited due to impurity. Thus, quality-controlled production of recombinant lectin is necessary. In this study, the algal lectin BPL3 (Bryopsis plumosa lectin 3) was successfully produced using a bacterial expression system, BL21(DE3), with an artificial repeated structure (dimeric construct). Recombinant dimeric BPL3 (rD2BPL3) was confirmed by LC-MS/MS spectrometry. Expression efficiency was greater for the construct with the repeat structure (rD2BPL3) than the monomeric form (rD1BPL3). Optimal conditions for expression were 1 mM IPTG at 20 °C. Recombinant lectin was purified under denaturing conditions and refolded by the flash dilution method. Recombinant BPL3 was solubilized in 1× PBS containing 2 M urea. rD2BPL3 showed strong hemagglutination activity using human erythrocyte. rD2BPL3 had a similar sugar specificity to that of the native protein, i.e., to N-acetyl-glucosamine (GlcNAc) and N-acetyl-galactosamine (GalNAc). Glycan array results showed that recombinant BPL3 and native BPL3 exhibited different binding properties. Both showed weak binding activity to α-Man-Sp. Native BPL3 showed strong binding specificity to the alpha conformation of amino sugars, and rD2BPL3 had binding activity to the beta conformation. The process developed in this study was suitable for the quality-controlled large-scale production of recombinant lectins. MDPI 2018-01-05 /pmc/articles/PMC5793061/ /pubmed/29303968 http://dx.doi.org/10.3390/md16010013 Text en © 2018 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 Hwang, Hyun-Ju Han, Jin-Woo Kim, Gwang Hoon Han, Jong Won Functional Expression and Characterization of the Recombinant N-Acetyl-Glucosamine/N-Acetyl-Galactosamine-Specific Marine Algal Lectin BPL3 |
title | Functional Expression and Characterization of the Recombinant N-Acetyl-Glucosamine/N-Acetyl-Galactosamine-Specific Marine Algal Lectin BPL3 |
title_full | Functional Expression and Characterization of the Recombinant N-Acetyl-Glucosamine/N-Acetyl-Galactosamine-Specific Marine Algal Lectin BPL3 |
title_fullStr | Functional Expression and Characterization of the Recombinant N-Acetyl-Glucosamine/N-Acetyl-Galactosamine-Specific Marine Algal Lectin BPL3 |
title_full_unstemmed | Functional Expression and Characterization of the Recombinant N-Acetyl-Glucosamine/N-Acetyl-Galactosamine-Specific Marine Algal Lectin BPL3 |
title_short | Functional Expression and Characterization of the Recombinant N-Acetyl-Glucosamine/N-Acetyl-Galactosamine-Specific Marine Algal Lectin BPL3 |
title_sort | functional expression and characterization of the recombinant n-acetyl-glucosamine/n-acetyl-galactosamine-specific marine algal lectin bpl3 |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5793061/ https://www.ncbi.nlm.nih.gov/pubmed/29303968 http://dx.doi.org/10.3390/md16010013 |
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