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Natural products isolation from Trichoderma reesei

Authors: Shenouda, Mary Labib Kamel Labib;

Natural products isolation from Trichoderma reesei

Abstract

The focus of the present work is to develop Trichoderma reesei as a heterologous host for the production of natural products and to explore the biosynthetic potential of this fungus by expressing one of its cryptic PKS-NRPS gene clusters. Developing T. reesei as a heterologous host was first done by knocking out two of the main genes from the sorbicillin biosynthetic gene cluster in the auxotrophic strain T. reesei QM6a Δtmus53 Δpyr4 to develop a strain with cleaner metabolic background. Afterwards, a new vector with native pdc promoter and pyr4 selection marker was constructed and used to express the polyketide synthase aspks1 under the activity of the native pdc promoter. The transformed T. reesei strains were able to produce the expected natural product 3-methylorcinaldehyde 40 on different media and types of waste materials such as orange peel, banana, kiwi and potato peel as well as barley straw. Quantification of the produced compound 40 on potato peel showed the ability of the strain to produce up to 128 mg·kg-1 dry weight potato peel. A second vector with two more native promoters (enolase and cDNA1 promoters) was constructed to allow the expression of more than one gene simultaneously in T. reesei. The new vector was then used to express the first two genes from the tenellin biosynthetic gene cluster, i.e. PKS-NRPS (encoded by tenS) and ER (encoded by tenC) under the activity of the pdc and cDNA1 promoters, respectively. T. reesei transformants expressing tenS and tenC showed the production of the expected compound pretenellin A 77 on different media. The new strain was also able to produce 77 on banana peel, but not on barley straw under the tested cultivation conditions. Bioinformatic analysis of the T. reesei genome revealed the presence of 11 PKS and two PKS-NRPS genes. Since no PKS-NRPS products have been previously reported from T. reesei, one of the two PKS-NRPS (TRIREDRAFT_58285) gene clusters in T. reesei was investigated. This was done first by overexpression of the transcription factor, which showed limited success due to the very low titre of the produced compound. Therefore, heterologous expression of this gene cluster was done in Aspergillus oryzae NSAR1, which confirmed that this gene cluster is responsible for the production of ilicicolin-H 72 in addition to many other intermediates and shunt products produced by the expression host.

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Keywords

Ilicicolin-H, Dewey Decimal Classification::500 | Naturwissenschaften::540 | Chemie, Naturstoffe, Biosynthese, Trichoderma reesei, heterologer Ausdruck, Heterologous expression, Biosynthesis, Natural product

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selected citations
These citations are derived from selected sources.
This is an alternative to the "Influence" indicator, which also reflects the overall/total impact of an article in the research community at large, based on the underlying citation network (diachronically).
BIP!Citations provided by BIP!
popularity
This indicator reflects the "current" impact/attention (the "hype") of an article in the research community at large, based on the underlying citation network.
BIP!Popularity provided by BIP!
influence
This indicator reflects the overall/total impact of an article in the research community at large, based on the underlying citation network (diachronically).
BIP!Influence provided by BIP!
impulse
This indicator reflects the initial momentum of an article directly after its publication, based on the underlying citation network.
BIP!Impulse provided by BIP!
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