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Immobilization of the Highly Active UDP-Glucose Pyrophosphorylase From Thermocrispum agreste Provides a Highly Efficient Biocatalyst for the Production of UDP-Glucose
Biocatalysis that produces economically interesting compounds can be carried out by using free enzymes or microbial cells. However, often the cell metabolism does not allow the overproduction or secretion of activated sugars and thus downstream processing of these sugars is complicated. Here enzyme...
Autores principales: | , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Frontiers Media S.A.
2020
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7343719/ https://www.ncbi.nlm.nih.gov/pubmed/32714915 http://dx.doi.org/10.3389/fbioe.2020.00740 |
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author | Kumpf, Antje Kowalczykiewicz, Daria Szymańska, Katarzyna Mehnert, Maria Bento, Isabel Łochowicz, Aleksandra Pollender, André Jarzȩbski, Andrzej Tischler, Dirk |
author_facet | Kumpf, Antje Kowalczykiewicz, Daria Szymańska, Katarzyna Mehnert, Maria Bento, Isabel Łochowicz, Aleksandra Pollender, André Jarzȩbski, Andrzej Tischler, Dirk |
author_sort | Kumpf, Antje |
collection | PubMed |
description | Biocatalysis that produces economically interesting compounds can be carried out by using free enzymes or microbial cells. However, often the cell metabolism does not allow the overproduction or secretion of activated sugars and thus downstream processing of these sugars is complicated. Here enzyme immobilization comes into focus in order to stabilize the enzyme as well as to make the overall process economically feasible. Besides a robust immobilization method, a highly active and stable enzyme is needed to efficiently produce the product of choice. Herein, we report on the identification, gene expression, biochemical characterization as well as immobilization of the uridine-5′-diphosphate-glucose (UDP-glucose) pyrophosphorylase originating from the thermostable soil actinobacterium Thermocrispum agreste DSM 44070 (TaGalU). The enzyme immobilization was performed on organically modified mesostructured cellular foams (MCF) via epoxy and amino group to provide a stable and active biocatalyst. The soluble and highly active TaGalU revealed a V(max) of 1698 U mg(–1) (uridine-5′-triphosphate, UTP) and a K(m) of 0.15 mM (UTP). The optimum reaction temperature was determined to be 50°C. TaGalU was stable at this temperature for up to 30 min with a maximum loss of activity of 65%. Interestingly, immobilized TaGalU was stable at 50°C for at least 120 min without a significant loss of activity, which makes this enzyme an interesting biocatalyst for the production of UDP-glucose. |
format | Online Article Text |
id | pubmed-7343719 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2020 |
publisher | Frontiers Media S.A. |
record_format | MEDLINE/PubMed |
spelling | pubmed-73437192020-07-25 Immobilization of the Highly Active UDP-Glucose Pyrophosphorylase From Thermocrispum agreste Provides a Highly Efficient Biocatalyst for the Production of UDP-Glucose Kumpf, Antje Kowalczykiewicz, Daria Szymańska, Katarzyna Mehnert, Maria Bento, Isabel Łochowicz, Aleksandra Pollender, André Jarzȩbski, Andrzej Tischler, Dirk Front Bioeng Biotechnol Bioengineering and Biotechnology Biocatalysis that produces economically interesting compounds can be carried out by using free enzymes or microbial cells. However, often the cell metabolism does not allow the overproduction or secretion of activated sugars and thus downstream processing of these sugars is complicated. Here enzyme immobilization comes into focus in order to stabilize the enzyme as well as to make the overall process economically feasible. Besides a robust immobilization method, a highly active and stable enzyme is needed to efficiently produce the product of choice. Herein, we report on the identification, gene expression, biochemical characterization as well as immobilization of the uridine-5′-diphosphate-glucose (UDP-glucose) pyrophosphorylase originating from the thermostable soil actinobacterium Thermocrispum agreste DSM 44070 (TaGalU). The enzyme immobilization was performed on organically modified mesostructured cellular foams (MCF) via epoxy and amino group to provide a stable and active biocatalyst. The soluble and highly active TaGalU revealed a V(max) of 1698 U mg(–1) (uridine-5′-triphosphate, UTP) and a K(m) of 0.15 mM (UTP). The optimum reaction temperature was determined to be 50°C. TaGalU was stable at this temperature for up to 30 min with a maximum loss of activity of 65%. Interestingly, immobilized TaGalU was stable at 50°C for at least 120 min without a significant loss of activity, which makes this enzyme an interesting biocatalyst for the production of UDP-glucose. Frontiers Media S.A. 2020-07-02 /pmc/articles/PMC7343719/ /pubmed/32714915 http://dx.doi.org/10.3389/fbioe.2020.00740 Text en Copyright © 2020 Kumpf, Kowalczykiewicz, Szymańska, Mehnert, Bento, Łochowicz, Pollender, Jarzȩbski and Tischler. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. |
spellingShingle | Bioengineering and Biotechnology Kumpf, Antje Kowalczykiewicz, Daria Szymańska, Katarzyna Mehnert, Maria Bento, Isabel Łochowicz, Aleksandra Pollender, André Jarzȩbski, Andrzej Tischler, Dirk Immobilization of the Highly Active UDP-Glucose Pyrophosphorylase From Thermocrispum agreste Provides a Highly Efficient Biocatalyst for the Production of UDP-Glucose |
title | Immobilization of the Highly Active UDP-Glucose Pyrophosphorylase From Thermocrispum agreste Provides a Highly Efficient Biocatalyst for the Production of UDP-Glucose |
title_full | Immobilization of the Highly Active UDP-Glucose Pyrophosphorylase From Thermocrispum agreste Provides a Highly Efficient Biocatalyst for the Production of UDP-Glucose |
title_fullStr | Immobilization of the Highly Active UDP-Glucose Pyrophosphorylase From Thermocrispum agreste Provides a Highly Efficient Biocatalyst for the Production of UDP-Glucose |
title_full_unstemmed | Immobilization of the Highly Active UDP-Glucose Pyrophosphorylase From Thermocrispum agreste Provides a Highly Efficient Biocatalyst for the Production of UDP-Glucose |
title_short | Immobilization of the Highly Active UDP-Glucose Pyrophosphorylase From Thermocrispum agreste Provides a Highly Efficient Biocatalyst for the Production of UDP-Glucose |
title_sort | immobilization of the highly active udp-glucose pyrophosphorylase from thermocrispum agreste provides a highly efficient biocatalyst for the production of udp-glucose |
topic | Bioengineering and Biotechnology |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7343719/ https://www.ncbi.nlm.nih.gov/pubmed/32714915 http://dx.doi.org/10.3389/fbioe.2020.00740 |
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