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Title: Heterologous gene expression and functional analysis of a type III polyketide synthase from Aspergillus niger NRRL 328

Abstract

Type III polyketide synthases (PKSs) catalyze the formation of pyrone- and resorcinol-types aromatic polyketides. The genomic analysis of the filamentous fungus Aspergillus niger NRRL 328 revealed that this strain has a putative gene (chr-8-2: 2978617–2979847) encoding a type III PKS, although its functions are unknown. In this study, for functional analysis of this putative type III PKS designated as An-CsyA, cloning and heterologous expression of the An-CsyA gene (An-csyA) in Escherichia coli were performed. Recombinant His-tagged An-CsyA was successfully expressed in E. coli BL21 (DE3), purified by Ni{sup 2+}-affinity chromatography, and used for in vitro assay. Tests on the substrate specificity of the His-tagged An-CsyA with myriad acyl-CoAs as starter substrates and malonyl-CoA as extender substrate showed that His-tagged An-CsyA accepted fatty acyl-CoAs (C2-C14) and produced triketide pyrones (C2-C14), tetraketide pyrones (C2-C10), and pentaketide resorcinols (C10-C14). Furthermore, acetoacetyl-CoA, malonyl-CoA, isobutyryl-CoA, and benzoyl-CoA were also accepted as starter substrates, and both of triketide pyrones and tetraketide pyrones were produced. It is noteworthy that the His-tagged An-CsyA produced polyketides from malonyl-CoA as starter and extender substrates and produced tetraketide pyrones from short-chain fatty acyl-CoAs as starter substrates. Therefore, this is the first report showing the functional properties of An-CsyA different from those of othermore » fungal type III PKSs. -- Highlights: •Type III PKS from Aspergillus niger NRRL 328, An-CsyA, was cloned and characterized. •An-CsyA produced triketide pyrones, tetraketide pyrones and pentaketide resorcinols. •Functional properties of An-CsyA differs from those of other fungal type III PKSs.« less

Authors:
; ;
Publication Date:
OSTI Identifier:
22598717
Resource Type:
Journal Article
Journal Name:
Biochemical and Biophysical Research Communications
Additional Journal Information:
Journal Volume: 473; Journal Issue: 4; Other Information: Copyright (c) 2016 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved.; Country of input: International Atomic Energy Agency (IAEA); Journal ID: ISSN 0006-291X
Country of Publication:
United States
Language:
English
Subject:
60 APPLIED LIFE SCIENCES; ASPERGILLUS; CARBON 10; CARBON 14; CHROMATOGRAPHY; CLONING; COBALT ARSENIDES; ESCHERICHIA COLI; FUNCTIONAL ANALYSIS; GENES; IN VITRO; LACTONES; NIGER; PYRONES; RESORCINOL; SUBSTRATES

Citation Formats

Kirimura, Kohtaro, Watanabe, Shotaro, and Kobayashi, Keiichi. Heterologous gene expression and functional analysis of a type III polyketide synthase from Aspergillus niger NRRL 328. United States: N. p., 2016. Web. doi:10.1016/J.BBRC.2016.04.023.
Kirimura, Kohtaro, Watanabe, Shotaro, & Kobayashi, Keiichi. Heterologous gene expression and functional analysis of a type III polyketide synthase from Aspergillus niger NRRL 328. United States. doi:10.1016/J.BBRC.2016.04.023.
Kirimura, Kohtaro, Watanabe, Shotaro, and Kobayashi, Keiichi. Fri . "Heterologous gene expression and functional analysis of a type III polyketide synthase from Aspergillus niger NRRL 328". United States. doi:10.1016/J.BBRC.2016.04.023.
@article{osti_22598717,
title = {Heterologous gene expression and functional analysis of a type III polyketide synthase from Aspergillus niger NRRL 328},
author = {Kirimura, Kohtaro and Watanabe, Shotaro and Kobayashi, Keiichi},
abstractNote = {Type III polyketide synthases (PKSs) catalyze the formation of pyrone- and resorcinol-types aromatic polyketides. The genomic analysis of the filamentous fungus Aspergillus niger NRRL 328 revealed that this strain has a putative gene (chr-8-2: 2978617–2979847) encoding a type III PKS, although its functions are unknown. In this study, for functional analysis of this putative type III PKS designated as An-CsyA, cloning and heterologous expression of the An-CsyA gene (An-csyA) in Escherichia coli were performed. Recombinant His-tagged An-CsyA was successfully expressed in E. coli BL21 (DE3), purified by Ni{sup 2+}-affinity chromatography, and used for in vitro assay. Tests on the substrate specificity of the His-tagged An-CsyA with myriad acyl-CoAs as starter substrates and malonyl-CoA as extender substrate showed that His-tagged An-CsyA accepted fatty acyl-CoAs (C2-C14) and produced triketide pyrones (C2-C14), tetraketide pyrones (C2-C10), and pentaketide resorcinols (C10-C14). Furthermore, acetoacetyl-CoA, malonyl-CoA, isobutyryl-CoA, and benzoyl-CoA were also accepted as starter substrates, and both of triketide pyrones and tetraketide pyrones were produced. It is noteworthy that the His-tagged An-CsyA produced polyketides from malonyl-CoA as starter and extender substrates and produced tetraketide pyrones from short-chain fatty acyl-CoAs as starter substrates. Therefore, this is the first report showing the functional properties of An-CsyA different from those of other fungal type III PKSs. -- Highlights: •Type III PKS from Aspergillus niger NRRL 328, An-CsyA, was cloned and characterized. •An-CsyA produced triketide pyrones, tetraketide pyrones and pentaketide resorcinols. •Functional properties of An-CsyA differs from those of other fungal type III PKSs.},
doi = {10.1016/J.BBRC.2016.04.023},
journal = {Biochemical and Biophysical Research Communications},
issn = {0006-291X},
number = 4,
volume = 473,
place = {United States},
year = {2016},
month = {5}
}