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Production of the polyketide 6-deoxyerythronolide B in the heterologous host Bacillus subtilis

Polyketides, such as erythromycin, are complex natural products with diverse therapeutic applications. They are synthesized by multi-modular megaenzymes, so-called polyketide synthases (PKSs). The macrolide core of erythromycin, 6-deoxyerythronolide B (6dEB), is produced by the deoxyerythronolide B...

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Autores principales: Kumpfmüller, Jana, Methling, Karen, Fang, Lei, Pfeifer, Blaine A., Lalk, Michael, Schweder, Thomas
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Springer Berlin Heidelberg 2015
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4717160/
https://www.ncbi.nlm.nih.gov/pubmed/26432460
http://dx.doi.org/10.1007/s00253-015-6990-6
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author Kumpfmüller, Jana
Methling, Karen
Fang, Lei
Pfeifer, Blaine A.
Lalk, Michael
Schweder, Thomas
author_facet Kumpfmüller, Jana
Methling, Karen
Fang, Lei
Pfeifer, Blaine A.
Lalk, Michael
Schweder, Thomas
author_sort Kumpfmüller, Jana
collection PubMed
description Polyketides, such as erythromycin, are complex natural products with diverse therapeutic applications. They are synthesized by multi-modular megaenzymes, so-called polyketide synthases (PKSs). The macrolide core of erythromycin, 6-deoxyerythronolide B (6dEB), is produced by the deoxyerythronolide B synthase (DEBS) that consists of three proteins each with a size of 330–370 kDa. We cloned and investigated the expression of the corresponding gene cluster from Saccharopolyspora erythraea, which comprises more than 30 kb, in Bacillus subtilis. It is shown that the DEBS genes are functionally expressed in B. subtilis when the native eryAI–III operon was separated into three individual expression cassettes with optimized ribosomal binding sites. A synthesis of 6dEB could be detected by using the acetoin-inducible acoA promoter and a fed-batch simulating EnBase-cultivation strategy. B. subtilis was capable of the secretion of 6dEB into the medium. In order to improve the 6dEB production, several genomic modifications of this production strain were tested. This included the knockout of the native secondary metabolite clusters of B. subtilis for the synthesis of surfactin (26 kb), bacillaene (76 kb), and plipastatin (38 kb). It is revealed that the deletion of the prpBD operon, responsible for propionyl-CoA utilization, resulted in a significant increase of the 6dEB product yield when exogenous propionate is provided. Although the presented B. subtilis 6dEB production strain is not competitive with established Escherichia coli 6dEB production strains, the results of this study indicate that B. subtilis is a suitable heterologous host for the secretory production of a complex polyketide. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s00253-015-6990-6) contains supplementary material, which is available to authorized users.
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spelling pubmed-47171602016-01-25 Production of the polyketide 6-deoxyerythronolide B in the heterologous host Bacillus subtilis Kumpfmüller, Jana Methling, Karen Fang, Lei Pfeifer, Blaine A. Lalk, Michael Schweder, Thomas Appl Microbiol Biotechnol Biotechnological Products and Process Engineering Polyketides, such as erythromycin, are complex natural products with diverse therapeutic applications. They are synthesized by multi-modular megaenzymes, so-called polyketide synthases (PKSs). The macrolide core of erythromycin, 6-deoxyerythronolide B (6dEB), is produced by the deoxyerythronolide B synthase (DEBS) that consists of three proteins each with a size of 330–370 kDa. We cloned and investigated the expression of the corresponding gene cluster from Saccharopolyspora erythraea, which comprises more than 30 kb, in Bacillus subtilis. It is shown that the DEBS genes are functionally expressed in B. subtilis when the native eryAI–III operon was separated into three individual expression cassettes with optimized ribosomal binding sites. A synthesis of 6dEB could be detected by using the acetoin-inducible acoA promoter and a fed-batch simulating EnBase-cultivation strategy. B. subtilis was capable of the secretion of 6dEB into the medium. In order to improve the 6dEB production, several genomic modifications of this production strain were tested. This included the knockout of the native secondary metabolite clusters of B. subtilis for the synthesis of surfactin (26 kb), bacillaene (76 kb), and plipastatin (38 kb). It is revealed that the deletion of the prpBD operon, responsible for propionyl-CoA utilization, resulted in a significant increase of the 6dEB product yield when exogenous propionate is provided. Although the presented B. subtilis 6dEB production strain is not competitive with established Escherichia coli 6dEB production strains, the results of this study indicate that B. subtilis is a suitable heterologous host for the secretory production of a complex polyketide. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s00253-015-6990-6) contains supplementary material, which is available to authorized users. Springer Berlin Heidelberg 2015-10-02 2016 /pmc/articles/PMC4717160/ /pubmed/26432460 http://dx.doi.org/10.1007/s00253-015-6990-6 Text en © The Author(s) 2015 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made.
spellingShingle Biotechnological Products and Process Engineering
Kumpfmüller, Jana
Methling, Karen
Fang, Lei
Pfeifer, Blaine A.
Lalk, Michael
Schweder, Thomas
Production of the polyketide 6-deoxyerythronolide B in the heterologous host Bacillus subtilis
title Production of the polyketide 6-deoxyerythronolide B in the heterologous host Bacillus subtilis
title_full Production of the polyketide 6-deoxyerythronolide B in the heterologous host Bacillus subtilis
title_fullStr Production of the polyketide 6-deoxyerythronolide B in the heterologous host Bacillus subtilis
title_full_unstemmed Production of the polyketide 6-deoxyerythronolide B in the heterologous host Bacillus subtilis
title_short Production of the polyketide 6-deoxyerythronolide B in the heterologous host Bacillus subtilis
title_sort production of the polyketide 6-deoxyerythronolide b in the heterologous host bacillus subtilis
topic Biotechnological Products and Process Engineering
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4717160/
https://www.ncbi.nlm.nih.gov/pubmed/26432460
http://dx.doi.org/10.1007/s00253-015-6990-6
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