Application of the C3-Methyltransferase StspM1 for the Synthesis of the Natural Pyrroloindole Motif.

Even though pyrroloindoles are widely present in natural products with different kinds of biological activities, their selective synthesis remains challenging with existing tools in organic chemistry, and there is furthermore a demand for stereoselective and mild methods to access this structural motif. Nature uses C3-methyltransferases to form the pyrroloindole framework, starting from the amino acid tryptophan. In the present study, the SAM-dependent methyltransferase StspM1 from Streptomyces sp. HPH0547 is used to build the pyrroloindole structural motif in tryptophan-based diketopiperazines (DKP). The substrate scope of the enzyme regarding different Trp-Trp-DKP isomers was investigated on an experimental and computational level. After further characterization and optimization of the methylation reaction with a design of experiment approach, a preparative scale reaction with the immobilized enzyme including a SAM regeneration system was performed to show the synthetic use of this biocatalytic tool to access the pyrroloindole structural motif.

Modular Approach for the Synthesis and Bioactivity Profiling of 8,8'-Biflavones.

In this work, we report the scalable and modular synthesis of a library of 55 monomeric and dimeric flavonoids including 14 8,8'-biflavones. The sterically demanding tetra-ortho-substituted axis of an acetophenone dimer key intermediate was constructed in a regioselective manner using Fe-mediated oxidative coupling. This step was systematically optimized and performed on up to multigram scale. The biological activities of this compound library were evaluated, including cytotoxicity against healthy and malignant human cell lines, antimicrobial activity against the apicomplexan parasite Toxoplasma gondii, and antioxidant capacity. A marked increase in activity for the 8,8'-dimeric structures compared to that of their monomeric counterparts was observed. Several biflavones were identified with high selectivity indices (low cytotoxicity and high antiprotozoal activity), showing that this class of natural products may serve as lead structures for further investigations.

Aerobic C-N Bond Formation through Enzymatic Nitroso-Ene-Type Reactions.

The biocatalytic oxidation of acylated hydroxylamines enables the direct and selective introduction of nitrogen functionalities by activation of allylic C-H bonds. Utilizing either laccases or an oxidase/peroxidase couple for the formal dehydrogenation of N-hydroxycarbamates and hydroxamic acids with air as the terminal oxidant, acylnitroso species are generated under particularly mild aqueous conditions. The reactive intermediates undergo C-N bond formation through an ene-type mechanism and provide high yields both in intramolecular and intermolecular enzymatic aminations. Investigations on different pathways of the two biocatalytic systems and labelling studies provide more insight into this unprecedented promiscuity of classical oxidoreductases as catalysts for nitroso-based transformations.

Optochemical Control of Bacterial Gene Expression: Novel Photocaged Compounds for Different Promoter Systems.

Photocaged compounds are applied for implementing precise, optochemical control of gene expression in bacteria. To broaden the scope of UV-light-responsive inducer molecules, six photocaged carbohydrates were synthesized and photochemically characterized, with the absorption exhibiting a red-shift. Their differing linkage through ether, carbonate, and carbamate bonds revealed that carbonate and carbamate bonds are convenient. Subsequently, those compounds were successfully applied in vivo for controlling gene expression in E. coli via blue light illumination. Furthermore, benzoate-based expression systems were subjected to light control by establishing a novel photocaged salicylic acid derivative. Besides its synthesis and in vitro characterization, we demonstrate the challenging choice of a suitable promoter system for light-controlled gene expression in E. coli. We illustrate various bottlenecks during both photocaged inducer synthesis and in vivo application and possibilities to overcome them. These findings pave the way towards novel caged inducer-dependent systems for wavelength-selective gene expression.