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Creating functional engineered variants of the single-module non-ribosomal peptide synthetase IndC by T domain exchange

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Creating functional engineered variants of the single-module non-ribosomal peptide synthetase IndC by T domain exchange. / Beer, Ralf; Herbst, Konrad; Ignatiadis, Nikolaos; Kats, Ilia; Adlung, Lorenz; Meyer, Hannah; Niopek, Dominik; Christiansen, Tania; Georgi, Fanny; Kurzawa, Nils; Meichsner, Johanna; Rabe, Sophie; Riedel, Anja; Sachs, Joshua; Schessner, Julia; Schmidt, Florian; Walch, Philipp; Niopek, Katharina; Heinemann, Tim; Eils, Roland; Di Ventura, Barbara.

In: MOL BIOSYST, Vol. 10, No. 7, 07.2014, p. 1709-18.

Research output: SCORING: Contribution to journal › SCORING: Journal article › Research › peer-review

Harvard

Beer, R, Herbst, K, Ignatiadis, N, Kats, I, Adlung, L, Meyer, H, Niopek, D, Christiansen, T, Georgi, F, Kurzawa, N, Meichsner, J, Rabe, S, Riedel, A, Sachs, J, Schessner, J, Schmidt, F, Walch, P, Niopek, K, Heinemann, T, Eils, R & Di Ventura, B 2014, 'Creating functional engineered variants of the single-module non-ribosomal peptide synthetase IndC by T domain exchange', MOL BIOSYST, vol. 10, no. 7, pp. 1709-18. https://doi.org/10.1039/c3mb70594c

APA

Beer, R., Herbst, K., Ignatiadis, N., Kats, I., Adlung, L., Meyer, H., Niopek, D., Christiansen, T., Georgi, F., Kurzawa, N., Meichsner, J., Rabe, S., Riedel, A., Sachs, J., Schessner, J., Schmidt, F., Walch, P., Niopek, K., Heinemann, T., ... Di Ventura, B. (2014). Creating functional engineered variants of the single-module non-ribosomal peptide synthetase IndC by T domain exchange. MOL BIOSYST, 10(7), 1709-18. https://doi.org/10.1039/c3mb70594c

Vancouver

Beer R, Herbst K, Ignatiadis N, Kats I, Adlung L, Meyer H et al. Creating functional engineered variants of the single-module non-ribosomal peptide synthetase IndC by T domain exchange. MOL BIOSYST. 2014 Jul;10(7):1709-18. https://doi.org/10.1039/c3mb70594c

Bibtex

@article{108024ff51bb4f9b8307f66cce786613,
title = "Creating functional engineered variants of the single-module non-ribosomal peptide synthetase IndC by T domain exchange",
abstract = "Non-ribosomal peptide synthetases (NRPSs) are enzymes that catalyze ribosome-independent production of small peptides, most of which are bioactive. NRPSs act as peptide assembly lines where individual, often interconnected modules each incorporate a specific amino acid into the nascent chain. The modules themselves consist of several domains that function in the activation, modification and condensation of the substrate. NRPSs are evidently modular, yet experimental proof of the ability to engineer desired permutations of domains and modules is still sought. Here, we use a synthetic-biology approach to create a small library of engineered NRPSs, in which the domain responsible for carrying the activated amino acid (T domain) is exchanged with natural or synthetic T domains. As a model system, we employ the single-module NRPS IndC from Photorhabdus luminescens that produces the blue pigment indigoidine. As chassis we use Escherichia coli. We demonstrate that heterologous T domain exchange is possible, even for T domains derived from different organisms. Interestingly, substitution of the native T domain with a synthetic one enhanced indigoidine production. Moreover, we show that selection of appropriate inter-domain linker regions is critical for functionality. Taken together, our results extend the engineering avenues for NRPSs, as they point out the possibility of combining domain sequences coming from different pathways, organisms or from conservation criteria. Moreover, our data suggest that NRPSs can be rationally engineered to control the level of production of the corresponding peptides. This could have important implications for industrial and medical applications.",
keywords = "Amino Acid Sequence, Bacterial Proteins/genetics, Escherichia coli/genetics, Genetic Variation, Peptide Synthases/genetics, Peptides/metabolism, Photorhabdus/enzymology, Piperidones/metabolism, Protein Engineering/methods, Sequence Homology, Amino Acid",
author = "Ralf Beer and Konrad Herbst and Nikolaos Ignatiadis and Ilia Kats and Lorenz Adlung and Hannah Meyer and Dominik Niopek and Tania Christiansen and Fanny Georgi and Nils Kurzawa and Johanna Meichsner and Sophie Rabe and Anja Riedel and Joshua Sachs and Julia Schessner and Florian Schmidt and Philipp Walch and Katharina Niopek and Tim Heinemann and Roland Eils and {Di Ventura}, Barbara",
year = "2014",
month = jul,
doi = "10.1039/c3mb70594c",
language = "English",
volume = "10",
pages = "1709--18",
journal = "MOL BIOSYST",
issn = "1742-206X",
publisher = "Royal Society of Chemistry",
number = "7",

}

RIS

TY - JOUR

T1 - Creating functional engineered variants of the single-module non-ribosomal peptide synthetase IndC by T domain exchange

AU - Beer, Ralf

AU - Herbst, Konrad

AU - Ignatiadis, Nikolaos

AU - Kats, Ilia

AU - Adlung, Lorenz

AU - Meyer, Hannah

AU - Niopek, Dominik

AU - Christiansen, Tania

AU - Georgi, Fanny

AU - Kurzawa, Nils

AU - Meichsner, Johanna

AU - Rabe, Sophie

AU - Riedel, Anja

AU - Sachs, Joshua

AU - Schessner, Julia

AU - Schmidt, Florian

AU - Walch, Philipp

AU - Niopek, Katharina

AU - Heinemann, Tim

AU - Eils, Roland

AU - Di Ventura, Barbara

PY - 2014/7

Y1 - 2014/7

N2 - Non-ribosomal peptide synthetases (NRPSs) are enzymes that catalyze ribosome-independent production of small peptides, most of which are bioactive. NRPSs act as peptide assembly lines where individual, often interconnected modules each incorporate a specific amino acid into the nascent chain. The modules themselves consist of several domains that function in the activation, modification and condensation of the substrate. NRPSs are evidently modular, yet experimental proof of the ability to engineer desired permutations of domains and modules is still sought. Here, we use a synthetic-biology approach to create a small library of engineered NRPSs, in which the domain responsible for carrying the activated amino acid (T domain) is exchanged with natural or synthetic T domains. As a model system, we employ the single-module NRPS IndC from Photorhabdus luminescens that produces the blue pigment indigoidine. As chassis we use Escherichia coli. We demonstrate that heterologous T domain exchange is possible, even for T domains derived from different organisms. Interestingly, substitution of the native T domain with a synthetic one enhanced indigoidine production. Moreover, we show that selection of appropriate inter-domain linker regions is critical for functionality. Taken together, our results extend the engineering avenues for NRPSs, as they point out the possibility of combining domain sequences coming from different pathways, organisms or from conservation criteria. Moreover, our data suggest that NRPSs can be rationally engineered to control the level of production of the corresponding peptides. This could have important implications for industrial and medical applications.

AB - Non-ribosomal peptide synthetases (NRPSs) are enzymes that catalyze ribosome-independent production of small peptides, most of which are bioactive. NRPSs act as peptide assembly lines where individual, often interconnected modules each incorporate a specific amino acid into the nascent chain. The modules themselves consist of several domains that function in the activation, modification and condensation of the substrate. NRPSs are evidently modular, yet experimental proof of the ability to engineer desired permutations of domains and modules is still sought. Here, we use a synthetic-biology approach to create a small library of engineered NRPSs, in which the domain responsible for carrying the activated amino acid (T domain) is exchanged with natural or synthetic T domains. As a model system, we employ the single-module NRPS IndC from Photorhabdus luminescens that produces the blue pigment indigoidine. As chassis we use Escherichia coli. We demonstrate that heterologous T domain exchange is possible, even for T domains derived from different organisms. Interestingly, substitution of the native T domain with a synthetic one enhanced indigoidine production. Moreover, we show that selection of appropriate inter-domain linker regions is critical for functionality. Taken together, our results extend the engineering avenues for NRPSs, as they point out the possibility of combining domain sequences coming from different pathways, organisms or from conservation criteria. Moreover, our data suggest that NRPSs can be rationally engineered to control the level of production of the corresponding peptides. This could have important implications for industrial and medical applications.

KW - Amino Acid Sequence

KW - Bacterial Proteins/genetics

KW - Escherichia coli/genetics

KW - Genetic Variation

KW - Peptide Synthases/genetics

KW - Peptides/metabolism

KW - Photorhabdus/enzymology

KW - Piperidones/metabolism

KW - Protein Engineering/methods

KW - Sequence Homology, Amino Acid

U2 - 10.1039/c3mb70594c

DO - 10.1039/c3mb70594c

M3 - SCORING: Journal article

C2 - 24457530

VL - 10

SP - 1709

EP - 1718

JO - MOL BIOSYST

JF - MOL BIOSYST

SN - 1742-206X

IS - 7

ER -