Dataset Information

Diversity-oriented combinatorial biosynthesis of benzenediol lactone scaffolds by subunit shuffling of fungal polyketide synthases.

ABSTRACT: Combinatorial biosynthesis aspires to exploit the promiscuity of microbial anabolic pathways to engineer the synthesis of new chemical entities. Fungal benzenediol lactone (BDL) polyketides are important pharmacophores with wide-ranging bioactivities, including heat shock response and immune system modulatory effects. Their biosynthesis on a pair of sequentially acting iterative polyketide synthases (iPKSs) offers a test case for the modularization of secondary metabolic pathways into "build-couple-pair" combinatorial synthetic schemes. Expression of random pairs of iPKS subunits from four BDL model systems in a yeast heterologous host created a diverse library of BDL congeners, including a polyketide with an unnatural skeleton and heat shock response-inducing activity. Pairwise heterocombinations of the iPKS subunits also helped to illuminate the innate, idiosyncratic programming of these enzymes. Even in combinatorial contexts, these biosynthetic programs remained largely unchanged, so that the iPKSs built their cognate biosynthons, coupled these building blocks into chimeric polyketide intermediates, and catalyzed intramolecular pairing to release macrocycles or ?-pyrones. However, some heterocombinations also provoked stuttering, i.e., the relaxation of iPKSs chain length control to assemble larger homologous products. The success of such a plug and play approach to biosynthesize novel chemical diversity bodes well for bioprospecting unnatural polyketides for drug discovery.

SUBMITTER: Xu Y

PROVIDER: S-EPMC4151742 | biostudies-literature | 2014 Aug

REPOSITORIES: biostudies-literature

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Diversity-oriented combinatorial biosynthesis of benzenediol lactone scaffolds by subunit shuffling of fungal polyketide synthases.

Xu Yuquan Y Zhou Tong T Zhang Shuwei S Espinosa-Artiles Patricia P Wang Luoyi L Zhang Wei W Lin Min M Gunatilaka A A Leslie AA Zhan Jixun J Molnár István I

Proceedings of the National Academy of Sciences of the United States of America 20140721 34

Combinatorial biosynthesis aspires to exploit the promiscuity of microbial anabolic pathways to engineer the synthesis of new chemical entities. Fungal benzenediol lactone (BDL) polyketides are important pharmacophores with wide-ranging bioactivities, including heat shock response and immune system modulatory effects. Their biosynthesis on a pair of sequentially acting iterative polyketide synthases (iPKSs) offers a test case for the modularization of secondary metabolic pathways into "build-cou ...[more]

PMID: 25049383

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Project description:The goal of this study was to develop a tool specifically designed to identify iterative polyketide synthases (iPKSs) from predicted fungal proteomes. A fungi-based PKS prediction model, specifically for fungal iPKSs, was developed using profile hidden Markov models (pHMMs) based on two essential iPKS domains, the β-ketoacyl synthase (KS) domain and acyltransferase (AT) domain, derived from fungal iPKSs. This fungi-based PKS prediction model was initially tested on the well-annotated proteome of Fusarium graminearum, identifying 15 iPKSs that matched previous predictions and gene disruption studies. These fungi-based pHMMs were subsequently applied to the predicted fungal proteomes of Alternaria brassicicola, Fusarium oxysporum f.sp. lycopersici, Verticillium albo-atrum and Verticillium dahliae. The iPKSs predicted were compared against those predicted by the currently available mixed-kingdom PKS models that include both bacterial and fungal sequences. These mixed-kingdom models have been proven previously by others to be better in predicting true iPKSs from non-iPKSs compared with other available models (e.g. Pfam and TIGRFAM). The fungi-based model was found to perform significantly better on fungal proteomes than the mixed-kingdom PKS model in accuracy, sensitivity, specificity and precision. In addition, the model was capable of predicting the reducing nature of fungal iPKSs by comparison of the bit scores obtained from two separate reducing and nonreducing pHMMs for each domain, which was confirmed by phylogenetic analysis of the KS domain. Biological confirmation of the predictions was obtained by polymerase chain reaction (PCR) amplification of the KS and AT domains of predicted iPKSs from V. dahliae using domain-specific primers and genomic DNA, followed by sequencing of the PCR products. It is expected that the fungi-based PKS model will prove to be a useful tool for the identification and annotation of fungal PKSs from predicted proteomes.

Dataset Information

Diversity-oriented combinatorial biosynthesis of benzenediol lactone scaffolds by subunit shuffling of fungal polyketide synthases.

Publications

Diversity-oriented combinatorial biosynthesis of benzenediol lactone scaffolds by subunit shuffling of fungal polyketide synthases.

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