Dissecting modular synthases through inhibition: A complementary chemical and genetic approach

Christopher R. Vickery; Ian P.  McCulloch; Eva Sonnenschein; Joris Beld; Joseph P. Noel; Michael D. Burkart

doi:10.1016/j.bmcl.2019.126820

Dissecting modular synthases through inhibition: A complementary chemical and genetic approach

Christopher R. Vickery, Ian P. McCulloch, Eva Sonnenschein, Joris Beld, Joseph P. Noel, Michael D. Burkart^*

^*Corresponding author for this work

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

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Abstract

Modular synthases, such as fatty acid, polyketide, and non-ribosomal peptide synthases (NRPSs), are sophisticated machineries essential in both primary and secondary metabolism. Various techniques have been developed to understand their genetic background and enzymatic abilities. However, uncovering the actual biosynthetic pathways remains challenging. Herein, we demonstrate a pipeline to study an assembly line synthase by interrogating the enzymatic function of each individual enzymatic domain of BpsA, a NRPS that produces the blue 3,3′-bipyridyl pigment indigoidine. Specific inhibitors for each biosynthetic domain of BpsA were obtained or synthesized, and the enzymatic performance of BpsA upon addition of each inhibitor was monitored by pigment development in vitro and in living bacteria. The results were verified using genetic mutants to inactivate each domain. Finally, the results complemented the currently proposed biosynthetic pathway of BpsA.

Original language	English
Article number	126820
Journal	Bioorganic & Medicinal Chemistry Letters
Volume	30
Issue number	2
Number of pages	6
ISSN	0960-894X
DOIs	https://doi.org/10.1016/j.bmcl.2019.126820
Publication status	Published - 2020

Keywords

Non-ribosomal peptide synthetase
Indigoidine
BpsA
Natural products
Domain-specific inhibitors

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title = "Dissecting modular synthases through inhibition: A complementary chemical and genetic approach",

abstract = "Modular synthases, such as fatty acid, polyketide, and non-ribosomal peptide synthases (NRPSs), are sophisticated machineries essential in both primary and secondary metabolism. Various techniques have been developed to understand their genetic background and enzymatic abilities. However, uncovering the actual biosynthetic pathways remains challenging. Herein, we demonstrate a pipeline to study an assembly line synthase by interrogating the enzymatic function of each individual enzymatic domain of BpsA, a NRPS that produces the blue 3,3′-bipyridyl pigment indigoidine. Specific inhibitors for each biosynthetic domain of BpsA were obtained or synthesized, and the enzymatic performance of BpsA upon addition of each inhibitor was monitored by pigment development in vitro and in living bacteria. The results were verified using genetic mutants to inactivate each domain. Finally, the results complemented the currently proposed biosynthetic pathway of BpsA.",

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AU - Vickery, Christopher R.

AU - McCulloch, Ian P.

AU - Sonnenschein, Eva

AU - Beld, Joris

AU - Noel, Joseph P.

AU - Burkart, Michael D.

PY - 2020

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N2 - Modular synthases, such as fatty acid, polyketide, and non-ribosomal peptide synthases (NRPSs), are sophisticated machineries essential in both primary and secondary metabolism. Various techniques have been developed to understand their genetic background and enzymatic abilities. However, uncovering the actual biosynthetic pathways remains challenging. Herein, we demonstrate a pipeline to study an assembly line synthase by interrogating the enzymatic function of each individual enzymatic domain of BpsA, a NRPS that produces the blue 3,3′-bipyridyl pigment indigoidine. Specific inhibitors for each biosynthetic domain of BpsA were obtained or synthesized, and the enzymatic performance of BpsA upon addition of each inhibitor was monitored by pigment development in vitro and in living bacteria. The results were verified using genetic mutants to inactivate each domain. Finally, the results complemented the currently proposed biosynthetic pathway of BpsA.

AB - Modular synthases, such as fatty acid, polyketide, and non-ribosomal peptide synthases (NRPSs), are sophisticated machineries essential in both primary and secondary metabolism. Various techniques have been developed to understand their genetic background and enzymatic abilities. However, uncovering the actual biosynthetic pathways remains challenging. Herein, we demonstrate a pipeline to study an assembly line synthase by interrogating the enzymatic function of each individual enzymatic domain of BpsA, a NRPS that produces the blue 3,3′-bipyridyl pigment indigoidine. Specific inhibitors for each biosynthetic domain of BpsA were obtained or synthesized, and the enzymatic performance of BpsA upon addition of each inhibitor was monitored by pigment development in vitro and in living bacteria. The results were verified using genetic mutants to inactivate each domain. Finally, the results complemented the currently proposed biosynthetic pathway of BpsA.

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KW - Natural products

KW - Domain-specific inhibitors

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Dissecting modular synthases through inhibition: A complementary chemical and genetic approach

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