Heterologous Biosynthesis of Kauralexin A1 in Saccharomyces cerevisiae through Metabolic and Enzyme Engineering.

Kauralexin A1 (KA1) is a key intermediate of the kauralexin A series metabolites of maize phytoalexins. However, their application is severely limited by their low abundance in maize. In this study, an efficient biosynthetic pathway was constructed to produce KA1 in Saccharomyces cerevisiae. Also, metabolic and enzyme engineering strategies were applied to construct the high-titer strains, such as chassis modification, screening synthases, the colocalization of enzymes, and multiple genomic integrations. First, the KA1 precursor ent-kaurene was synthesized using the efficient diterpene synthase GfCPS/KS from Fusarium fujikuroi, and optimized to reach 244.36 mg/L in shake flasks, which displayed a 200-fold increase compared to the initial strain. Then, the KA1 was produced under the catalysis of ZmCYP71Z18 from Zea mays and SmCPR1 from Salvia miltiorrhiza, and the titer was further improved by integrating the fusion protein into the genome. Finally, an ent-kaurene titer of 763.23 mg/L and a KA1 titer of 42.22 mg/L were achieved through a single-stage fed-batch fermentation in a 5 L bioreactor. This is the first report of the heterologous biosynthesis of maize diterpene phytoalexins in S. cerevisiae, which lays a foundation for further pathway reconstruction and biosynthesis of the kauralexin A series maize phytoalexins.

Vinylene-Linked Covalent Organic Frameworks (COFs) with Symmetry-Tuned Polarity and Photocatalytic Activity.

The polarity of a semiconducting molecule affects its intrinsic photophysical properties, which can be tuned by varying the molecular geometry. Herein, we developed a D3h -symmetric tricyanomesitylene as a new monomer which could be reticulated into a vinylene-linked covalent organic framework (g-C54 N6 -COF) via Knoevenagel condensation with another D3h -symmetric monomer 2,4,6-tris(4'-formyl-biphenyl-4-yl)-1,3,5-triazine. Replacing tricyanomesitylene with a C2v -symmetric 3,5-dicyano-2,4,6-trimethylpyridine gave a less-symmetric vinylene-linked COF (g-C52 N6 -COF). The octupolar conjugated characters of g-C54 N6 -COF were reflected in its scarce solvatochromic effects either in ground or excited states, and endowed it with more promising semiconducting behavior as compared with g-C52 N6 -COF, such as enhanced light-harvesting and excellent photo-induced charge generation and separation. Along with the matched energy level, g-C54 N6 -COF enabled the two-half reactions of photocatalytic water splitting with an average O2 evolution rate of 51.0 µmol h-1 g-1 and H2 evolution rate of 2518.9 µmol h-1 g-1 . Such values are among the highest of state-of-the-art COF photocatalysts.

Vinylene-Bridged Two-Dimensional Covalent Organic Frameworks via Knoevenagel Condensation of Tricyanomesitylene.

Vinylene-bridged covalent organic frameworks (COFs) have shown great potential for advanced applications because of their high chemical stability and intriguing semiconducting properties. Exploring new functional monomers available for the reticulation of vinylene-bridged COFs and establishing effective reaction conditions are extremely desired for enlarging the realm of this kind of material. In this work, a series of vinylene-bridged two-dimensional (2D) COFs are synthesized by Knoevenagel condensation of tricyanomesitylene with ditopic or tritopic aromatic aldehydes. With use of appropriate secondary amines as catalysts, high-crystalline vinylene-bridged COFs were achieved, exhibiting long-range ordered structures, well-defined nanochannels, high surface areas (up to 1231 m2 g-1), and excellent photophysical properties. Under a low loading amount and short reaction time, they enable aerobic photocatalytic transformation of arylboronic acids to phenols with high efficiency and excellent recyclability. This work demonstrates a new functional monomer, tricyanomesitylene, feasible for the general synthesis of vinylene-bridged COFs with potential application in photocatalytic organic transformation, which instigates further research on such kind of material.

An Olefin-Linked Covalent Organic Framework as a Flexible Thin-Film Electrode for a High-Performance Micro-Supercapacitor.

Developing effective synthetic strategies as well as enriching functionalities for sp2 -carbon-linked covalent organic frameworks (COFs) still remains a challenge. Now, taking advantage of a variant of Knoevenagel condensation, a new fully conjugated COF (g-C34 N6 -COF) linked by unsubstituted C=C bonds was synthesized. Integrating 3,5-dicyano-2,4,6-trimethylpyridine and 1,3,5-triazine units into the molecular framework leads to the enhanced π-electron communication and electrochemical activity. This COF shows uniform nanofibrous morphology. By assembling it with carbon nanotubes, a flexible thin-film electrode for a micro-supercapacitor (MSC) can be easily obtained. The resultant COF-based MSC shows an areal capacitance of up to 15.2 mF cm-2 , a high energy density of up to 7.3 mWh cm-3 , and remarkable rate capability. These values are among the highest for state-of-the-art MSCs. Moreover, this device exhibits excellent flexibility and integration capability.

Two-dimensional semiconducting covalent organic frameworks via condensation at arylmethyl carbon atoms.

Construction of organic semiconducting materials with in-plane π-conjugated structures and robustness through carbon-carbon bond linkages, alternatively as organic graphene analogs, is extremely desired for powerfully optoelectrical conversion. However, the poor reversibility for sp2 carbon bond forming reactions makes them unavailable for building high crystalline well-defined organic structures through a self-healing process, such as covalent organic frameworks (COFs). Here we report a scalable solution-processing approach to synthesize a family of two-dimensional (2D) COFs with trans-disubstituted C = C linkages via condensation reaction at arylmethyl carbon atoms on the basis of 3,5-dicyano-2,4,6-trimethylpyridine and linear/trigonal aldehyde (i.e., 4,4″-diformyl-p-terphenyl, 4,4'-diformyl-1,1'-biphenyl, or 1,3,5-tris(4-formylphenyl)benzene) monomers. Such sp2 carbon-jointed-pyridinyl frameworks, featuring crystalline honeycomb-like structures with high surface areas, enable driving two half-reactions of water splitting separately under visible light irradiation, comparable to graphitic carbon nitride (g-C3N4) derivatives.