From the Total Synthesis of Semi-Viriditoxin, Semi-Viriditoxic Acid and Dimeric Naphthopyranones to their Biological Activities in Burkitt B Cell Lymphoma.

Dimeric naphthopyranones are known to be biologically active, however, for the corresponding monomeric naphthopyranones this information is still elusive. Here the first enantioselective total synthesis of semi-viriditoxic acid as well as the synthesis of semi-viriditoxin and derivatives is reported. The key intermediate in the synthesis of naphthopyranones is an α,ß-unsaturated δ-lactone, which we synthesized in two different ways (Ghosez-cyclization and Grubbs ring-closing metathesis), while the domino-Michael-Dieckmann reaction of the α,ß-unsaturated δ-lactone with an orsellinic acid derivative is the key reaction. A structure-activity relationship study was performed measuring the cytotoxicity in Burkitt B lymphoma cells (Ramos). The dimeric structure was found to be crucial for biological activity: Only the dimeric naphthopyranones showed cytotoxic and apoptotic activity, whereas the monomers did not display any activity at all.

Biocatalytic C3-Indole Methylation-A Useful Tool for the Natural-Product-Inspired Stereoselective Synthesis of Pyrroloindoles.

Enantioselective synthesis of bioactive compounds bearing a pyrroloindole framework is often laborious. In contrast, there are several S-adenosyl methionine (SAM)-dependent methyl transferases known for stereo- and regioselective methylation at the C3 position of various indoles, directly leading to the formation of the desired pyrroloindole moiety. Herein, the SAM-dependent methyl transferase PsmD from Streptomyces griseofuscus, a key enzyme in the biosynthesis of physostigmine, is characterized in detail. The biochemical properties of PsmD and its substrate scope were demonstrated. Preparative scale enzymatic methylation including SAM regeneration was achieved for three selected substrates after a design-of-experiment optimization.

First enantioselective total synthesis of altersolanol A.

The first enantioselective total synthesis of altersolanol A, a secondary metabolite from the endophytic fungi Stemphylium globuliferum and Alternaria solani, is described. The key step towards the tetrahydroanthraquinone core was an asymmetric Diels-Alder (D-A) cycloaddition promoted by (R)-3,3'-diphenyl-BINOL/boron Lewis acid with good to excellent yields and excellent diastereo- and enantioselectivity (>95 : 5 dr and 98 : 2 er).

Redesigning Aldolase Stereoselectivity by Homologous Grafting.

The 2-deoxy-d-ribose-5-phosphate aldolase (DERA) offers access to highly desirable building blocks for organic synthesis by catalyzing a stereoselective C-C bond formation between acetaldehyde and certain electrophilic aldehydes. DERA´s potential is particularly highlighted by the ability to catalyze sequential, highly enantioselective aldol reactions. However, its synthetic use is limited by the absence of an enantiocomplementary enzyme. Here, we introduce the concept of homologous grafting to identify stereoselectivity-determining amino acid positions in DERA. We identified such positions by structural analysis of the homologous aldolases 2-keto-3-deoxy-6-phosphogluconate aldolase (KDPG) and the enantiocomplementary enzyme 2-keto-3-deoxy-6-phosphogalactonate aldolase (KDPGal). Mutation of these positions led to a slightly inversed enantiopreference of both aldolases to the same extent. By transferring these sequence motifs onto DERA we achieved the intended change in enantioselectivity.

Enantioselective allylic hydroxylation of ω-alkenoic acids and esters by P450 BM3 monooxygenase.

Chiral allylic alcohols of ω-alkenoic acids and derivatives thereof are highly important building blocks for the synthesis of biologically active compounds. The direct enantioselective C-H oxidation of linear terminal olefins offers the shortest route toward these compounds, but known synthetic methods are limited and suffer from low selectivities. Described herein is an enzymatic approach using the P450 BM3 monooxygenase mutant A74G/L188Q, which catalyzes allylic hydroxylation with high to excellent chemo- and enantioselectivities providing the desirable secondary alcohols.

Chemo-enzymatic modification of poly-N-acetyllactosamine (LacNAc) oligomers and N,N-diacetyllactosamine (LacDiNAc) based on galactose oxidase treatment.

The importance of glycans in biological systems is highlighted by their various functions in physiological and pathological processes. Many glycan epitopes on glycoproteins and glycolipids are based on N-acetyllactosamine units (LacNAc; Galß1,4GlcNAc) and often present on extended poly-LacNAc glycans ([Galß1,4GlcNAc](n)). Poly-LacNAc itself has been identified as a binding motif of galectins, an important class of lectins with functions in immune response and tumorigenesis. Therefore, the synthesis of natural and modified poly-LacNAc glycans is of specific interest for binding studies with galectins as well as for studies of their possible therapeutic applications. We present the oxidation by galactose oxidase and subsequent chemical or enzymatic modification of terminal galactose and N-acetylgalactosamine residues of poly-N-acetyllactosamine (poly-LacNAc) oligomers and N,N-diacetyllactosamine (LacDiNAc) by galactose oxidase. Product formation starting from different poly-LacNAc oligomers was characterised and optimised regarding formation of the C6-aldo product. Further modification of the aldehyde containing glycans, either by chemical conversion or enzymatic elongation, was established. Base-catalysed ß-elimination, coupling of biotin-hydrazide with subsequent reduction to the corresponding hydrazine linkage, and coupling by reductive amination to an amino-functionalised poly-LacNAc oligomer were performed and the products characterised by LC-MS and NMR analysis. Remarkably, elongation of terminally oxidised poly-LacNAc glycans by ß3GlcNAc- and ß4Gal-transferase was also successful. In this way, a set of novel, modified poly-LacNAc oligomers containing terminally and/or internally modified galactose residues were obtained, which can be used for binding studies and various other applications.

Enzymatic synthesis of dimeric glycomimetic ligands of NK cell activation receptors.

This work reveals new structural relationships in the complex process of the interaction between activation receptors of natural killer cells (rat NKR-P1, human CD69) and novel bivalent carbohydrate glycomimetics. The length, glycosylation pattern and linker structure of receptor ligands were examined with respect to their ability to precipitate the receptor protein from solution, which simulates the in vivo process of receptor aggregation during NK cell activation. It was found that di-LacdiNAc triazole compounds show optimal performance, reaching up to 100% precipitation of the present protein receptors, and achieving high immunostimulatory activities without any tendency to trigger activation-induced apoptosis. In the synthesis of the compounds tested, two enzymatic approaches were applied. Whereas a ß-N-acetylhexosaminidase could only glycosylate one of the two acceptor sites available with yields below 10%, the Y284L mutant of human placental ß1,4-galactosyltransferase-1 worked as a perfect synthetic tool, accomplishing even quantitative glycosylation at both acceptor sites and with absolute regioselectivity for the C-4 position. This work insinuates new directions for further ligand structure optimisation and demonstrates the strong synthetic potential of the mutant human placental ß1,4-galactosyltransferase-1 in the synthesis of multivalent glycomimetics and glycomaterials.