Browsing by Author "Wang, Clay C. C."

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Engineering Fungal Nonreducing Polyketide Synthase by Heterologous Expression and Domain Swapping
(2013-02-15)
Yeh, Hsu-Hua
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Chang, Shu-Lin
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Chiang, Yi-Ming
;
Bruno, Kenneth S.
;
Oakley, Berl R.
We reannotated the A. niger NR-PKS gene, e_gw1_19.204, and its downstream R domain gene, est_GWPlus_C_190476, as a single gene which we named dtbA. Heterologous expression of dtbA In A. nidulans demonstrated that DtbA protein produces two polyketides, 2,4-dihydroxy-3,5,6-trimethylbenzaldehyde (1) and 6-ethyl-2,4-dihydroxy-3,5-dimethylbenzaldehyde (2). Generation of DtbA Delta R+TE chimeric PKSs by swapping the DtbA R domain with the AusA (austinol biosynthesis) or ANID_06448 TE domain enabled the production of two metabolites with carboxylic acids replacing the corresponding aldehydes.
Illuminating the Diversity of Aromatic Polyketide Synthases in Aspergillus nidulans
(2012-05-16)
Ahuja, Manmeet
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Chiang, Yi-Ming
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Chang, Shu-Lin
;
Praseuth, Mike B.
;
Entwistle, Ruth
Genome sequencing has revealed that fungi have the ability to synthesize many more natural products (NPs) than are currently known, but methods for obtaining suitable expression of NPs have been inadequate. We have developed a successful strategy that bypasses normal regulatory mechanisms. By efficient gene targeting, we have replaced, en masse, the promoters of nonreducing polyketide synthase (NR-PKS) genes, key genes in NP biosynthetic pathways, and other genes necessary for NR-PKS product formation or release. This has allowed us to determine the products of eight NR-PKSs of Aspergillus nidulans, including seven novel compounds, as well as the NR-PKS genes required for the synthesis of the toxins alternariol (8) and cichorine (19).
Molecular genetic analysis reveals that a nonribosomal peptide synthetase-like (NRPS-like) gene in Aspergillus nidulans is responsible for microperfuranone biosynthesis
(2012-11-01)
Yeh, Hsu-Hua
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Chiang, Yi-Ming
;
Entwistle, Ruth
;
Ahuja, Manmeet
;
Lee, Kuan-Han
Genome sequencing of Aspergillus species including Aspergillus nidulans has revealed that there are far more secondary metabolite biosynthetic gene clusters than secondary metabolites isolated from these organisms. This implies that these organisms can produce additional secondary metabolites, which have not yet been elucidated. The A. nidulans genome contains 12 nonribosomal peptide synthetase (NRPS), one hybrid polyketide synthase/NRPS, and 14 NRPS-like genes. The only NRPS-like gene in A. nidulans with a known product is tdiA, which is involved in terrequinone A biosynthesis. To attempt to identify the products of these NRPS-like genes, we replaced the native promoters of the NRPS-like genes with the inducible alcohol dehydrogenase (alcA) promoter. Our results demonstrated that induction of the single NRPS-like gene AN3396.4 led to the enhanced production of microperfuranone. Furthermore, heterologous expression of AN3396.4 in Aspergillus niger confirmed that only one NRPS-like gene, AN3396.4, is necessary for the production of microperfuranone.
Reconstitution of the early steps of gliotoxin biosynthesis in Aspergillus nidulans reveals the role of the monooxygenase GliC
(2013-04-01)
Chang, Shu-Lin
;
Chiang, Yi-Ming
;
Yeh, Hsu-Hua
;
Wu, Tung-Kung
;
Wang, Clay C. C.
The gliotoxin, a member of the epipolythiodioxopiperazine (ETP), has received considerable attention from the scientific community for its wide range of biological activity. Despite the identification of gliotoxin cluster, however, the sequence of steps in the gliotoxin biosynthesis has remained elusive. As an alternative to the gene knock-out and biochemical approaches used so far, here we report using a heterologous expression approach to determine the sequence of the early steps of gliotoxin biosynthesis in Aspergillus nidulans. We identified the GliC, a monooxygenases that involved in the second step of gliotoxin biosynthesis pathway through the catalyzing the hydroxylation at the alpha-position of L-Phe. (C) 2013 Elsevier Ltd. All rights reserved.