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Creators/Authors contains: "Tang, Yi"

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  1. Abstract  

    Monoterpene indole alkaloids (MIAs) are a class of natural products comprised of thousands of structurally unique bioactive compounds with significant therapeutic values. Due to difficulties associated with isolation from native plant species and organic synthesis of these structurally complex molecules, microbial production of MIAs using engineered hosts are highly desired. In this work, we report the engineering of fully integrated Saccharomyces cerevisiae strains that allow de novo access to strictosidine, the universal precursor to thousands of MIAs at 30–40 mg/L. The optimization efforts were based on a previously reported yeast strain that is engineered to produce high titers of the monoterpene precursor geraniol through compartmentalization of mevalonate pathway in the mitochondria. Our approaches here included the use of CRISPR-dCas9 interference to identify mitochondria diphosphate transporters that negatively impact the titer of the monoterpene, followed by genetic inactivation; the overexpression of transcriptional regulators that increase cellular respiration and mitochondria biogenesis. Strain construction included the strategic integration of genes encoding both MIA biosynthetic and accessory enzymes into the genome under a variety of constitutive and inducible promoters. Following successful de novo production of strictosidine, complex alkaloids belonging to heteroyohimbine and corynantheine families were reconstituted in the host with introduction of additional downstream enzymes. We demonstrate that the serpentine/alstonine pair can be produced at ∼5 mg/L titer, while corynantheidine, the precursor to mitragynine can be produced at ∼1 mg/L titer. Feeding of halogenated tryptamine led to the biosynthesis of analogs of alkaloids in both families. Collectively, our yeast strain represents an excellent starting point to further engineer biosynthetic bottlenecks in this pathway and to access additional MIAs and analogs through microbial fermentation.

    One Sentence Summary

    An Saccharomyces cerevisiae-based microbial platform was developed for the biosynthesis of monoterpene indole alkaloids, including the universal precursor strictosidine and further modified heteroyohimbine and corynantheidine alkaloids.

     
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  2. Abstract Self-assembled systems have recently attracted extensive attention because they can display a wide range of phase morphologies in nanocomposites, providing a new arena to explore novel phenomena. Among these morphologies, a bicontinuous structure is highly desirable based on its high interface-to-volume ratio and 3D interconnectivity. A bicontinuous nickel oxide (NiO) and tin dioxide (SnO 2 ) heteroepitaxial nanocomposite is revealed here. By controlling their concentration, we fabricated tuneable self-assembled nanostructures from pillars to bicontinuous structures, as evidenced by TEM-energy-dispersive X-ray spectroscopy with a tortuous compositional distribution. The experimentally observed growth modes are consistent with predictions by first-principles calculations. Phase-field simulations are performed to understand 3D microstructure formation and extract key thermodynamic parameters for predicting microstructure morphologies in SnO 2 :NiO nanocomposites of other concentrations. Furthermore, we demonstrate significantly enhanced photovoltaic properties in a bicontinuous SnO 2 :NiO nanocomposite macroscopically and microscopically. This research shows a pathway to developing innovative solar cell and photodetector devices based on self-assembled oxides. 
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    Free, publicly-accessible full text available December 1, 2024
  3. Abstract

    Siderophores belonging to the ferrichrome family are essential for the viability of fungal species and play a key role for virulence of numerous pathogenic fungi. Despite their biological significance, our understanding of how these iron-chelating cyclic hexapeptides are assembled by non-ribosomal peptide synthetase (NRPS) enzymes remains poorly understood, primarily due to the nonlinearity exhibited by the domain architecture. Herein, we report the biochemical characterization of the SidC NRPS, responsible for construction of the intracellular siderophore ferricrocin. In vitro reconstitution of purified SidC reveals its ability to produce ferricrocin and its structural variant, ferrichrome. Application of intact protein mass spectrometry uncovers several non-canonical events during peptidyl siderophore biosynthesis, including inter-modular loading of amino acid substrates and an adenylation domain capable of poly-amide bond formation. This work expands the scope of NRPS programming, allows biosynthetic assignment of ferrichrome NRPSs, and sets the stage for reprogramming towards novel hydroxamate scaffolds.

     
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  4. Abstract

    We uncovered and reconstituted a concise biosynthetic pathway of the strained dipeptide (+)‐azonazine A from marine‐derivedAspergillus insulicola. Formation of the hexacyclic benzofuranoindoline ring system from cyclo‐(l‐Trp‐N‐methyl‐l‐Tyr) is catalyzed by a P450 enzyme through an oxidative cyclization. Supplementing the producing strain with various indole‐substituted tryptophan derivatives resulted in the generation of a series of azonazine A analogs.

     
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  5. Abstract

    We uncovered and reconstituted a concise biosynthetic pathway of the strained dipeptide (+)‐azonazine A from marine‐derivedAspergillus insulicola. Formation of the hexacyclic benzofuranoindoline ring system from cyclo‐(l‐Trp‐N‐methyl‐l‐Tyr) is catalyzed by a P450 enzyme through an oxidative cyclization. Supplementing the producing strain with various indole‐substituted tryptophan derivatives resulted in the generation of a series of azonazine A analogs.

     
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