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Author Kirimura, Kohtaro ♦ Watanabe, Shotaro ♦ Kobayashi, Keiichi
Source United States Department of Energy Office of Scientific and Technical Information
Content type Text
Language English
Subject Keyword APPLIED LIFE SCIENCES ♦ ASPERGILLUS ♦ CARBON 10 ♦ CARBON 14 ♦ CHROMATOGRAPHY ♦ CLONING ♦ COBALT ARSENIDES ♦ ESCHERICHIA COLI ♦ FUNCTIONAL ANALYSIS ♦ GENES ♦ IN VITRO ♦ LACTONES ♦ NIGER ♦ PYRONES ♦ RESORCINOL ♦ SUBSTRATES
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 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.
ISSN 0006291X
Educational Use Research
Learning Resource Type Article
Publisher Date 2016-05-13
Publisher Place United States
Journal Biochemical and Biophysical Research Communications
Volume Number 473
Issue Number 4


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