Engineered polyketide biosynthesis and biocatalysis in Escherichia coli

Polyketides are important bioactive natural products biosynthesized by bacteria, fungi, and plants. The enzymes that synthesize polyketides are collectively referred to as polyketide synthases (PKSs). Because many of the natural hosts that produce polyketides are difficult to culture or manipulate, establishing a universal heterologous host that is genetically tractable has become an important goal toward the engineered biosynthesis of polyketides and analogues. Here, we summarize the recent progresses in engineering Escherichia coli as a heterologous host for reconstituting PKSs of different types. Our increased understanding of PKS enzymology and structural biology, combined with new tools in protein engineering, metabolic engineering, and synthetic biology, has firmly established E. coli as a powerful host for producing polyketides

Light regulation of metabolic pathways in fungi

Light represents a major carrier of information in nature. The molecular machineries translating its electromagnetic energy (photons) into the chemical language of cells transmit vital signals for adjustment of virtually every living organism to its habitat. Fungi react to illumination in various ways, and we found that they initiate considerable adaptations in their metabolic pathways upon growth in light or after perception of a light pulse. Alterations in response to light have predominantly been observed in carotenoid metabolism, polysaccharide and carbohydrate metabolism, fatty acid metabolism, nucleotide and nucleoside metabolism, and in regulation of production of secondary metabolites. Transcription of genes is initiated within minutes, abundance and activity of metabolic enzymes are adjusted, and subsequently, levels of metabolites are altered to cope with the harmful effects of light or to prepare for reproduction, which is dependent on light in many cases. This review aims to give an overview on metabolic pathways impacted by light and to illustrate the physiological significance of light for fungi. We provide a basis for assessment whether a given metabolic pathway might be subject to regulation by light and how these properties can be exploited for improvement of biotechnological processes

Agrobacterium-mediated transformation as a tool for functional genomics in fungi.

Item does not contain fulltextIn the era of functional genomics, the need for tools to perform large-scale targeted and random mutagenesis is increasing. A potential tool is Agrobacterium-mediated fungal transformation. A. tumefaciens is able to transfer a part of its DNA (transferred DNA; T-DNA) to a wide variety of fungi and the number of fungi that can be transformed by Agrobacterium-mediated transformation (AMT) is still increasing. AMT has especially opened the field of molecular genetics for fungi that were difficult to transform with traditional methods or for which the traditional protocols failed to yield stable DNA integration. Because of the simplicity and efficiency of transformation via A. tumefaciens, it is relatively easy to generate a large number of stable transformants. In combination with the finding that the T-DNA integrates randomly and predominantly as a single copy, AMT is well suited to perform insertional mutagenesis in fungi. In addition, in various gene-targeting experiments, high homologous recombination frequencies were obtained, indicating that the T-DNA is also a useful substrate for targeted mutagenesis. In this review, we discuss the potential of the Agrobacterium DNA transfer system to be used as a tool for targeted and random mutagenesis in fungi