Total synthesis of new microbial sphingolipid-type signaling molecules

Sphingolipids are found in mammalian and bacterial membranes where they exert multiple intra- and intercellular functions. Therefore, binding partners of sphingolipids provide excellent targets for therapeutic approaches. Structurally related sulfonolipids (RIFs) are also of great interest for the interspecific interaction of Salpingoeca rosetta. In the present work, new access routes to both classes of compounds are demonstrated by the means of total chemical synthesis. Using a hydrozirconation/transmetallation reaction and a three-step one-pot reaction protection/deprotection sequence, several unnatural 2S,3S-6-hydroxy-4E-sphingenin-containing sphingolipids were prepared in only three (protected) or four (fully deprotected) consecutive synthetic steps. Among them is a fluorescently labeled derivative suitable for future biological studies. Furthermore, the first total synthesis of two RIF-2 diastereomers and one congener is reported, using 15 and eight synthetic steps, respectively. A decarboxylative cross-coupling reaction is used to synthesize the required branched alpha-hydroxy fatty acids. Furthermore, three different strategies are discussed in terms of their suitability for the synthesis of sulfonolipids: 1) the "cysteine" strategy, which uses cysteine as the central building block; 2) the "thioligation" strategy, which provides for a reaction similar to native chemical ligation to generate the peptide bond; and 3) the "late stage" strategy, which provides for the introduction of sulfur at the end of the total synthesis. Finally, the synthetic compounds are compared to the natural isolates in terms of their properties

Photo-labile BODIPY protecting groups for glycan synthesis

Protective groups that can be selectively removed under mild conditions are an essential aspect of carbohydrate chemistry. Groups that can be selectively removed by visible light are particularly attractive because carbohydrates are transparent to visible light. Here, different BODIPY protecting groups were explored for their utility during glycan synthesis. A BODIPY group bearing a boron difluoride unit is stable during glycosylations but can be cleaved with green light as illustrated by the assembly of a trisaccharide

Photo-labile BODIPY protecting groups for glycan synthesis

Protective groups that can be selectively removed under mild conditions are an essential aspect of carbohydrate chemistry. Groups that can be selectively removed by visible light are particularly attractive because carbohydrates are transparent to visible light. Here, different BODIPY protecting groups were explored for their utility during glycan synthesis. A BODIPY group bearing a boron difluoride unit is stable during glycosylations but can be cleaved with green light as illustrated by the assembly of a trisaccharide

Microwave-assisted automated glycan assembly

Automated synthesis of DNA, RNA, and peptides provides quickly and reliably important tools for biomedical research. Automated glycan assembly (AGA) is significantly more challenging as highly branched carbohydrates require strict regio- and stereocontrol during synthesis. A new AGA synthesizer enables rapid temperature adjustment from -40 °C to +100 °C to control glycosylations at low temperature and accelerates capping, protecting group removal, and glycan modifications by using elevated temperatures. Thereby, the temporary protecting group portfolio is extended from two to four orthogonal groups that give rise to oligosaccharides with up to four branches. In addition, sulfated glycans and unprotected glycans can be prepared. The new design reduces the typical coupling cycles from 100 min to 60 min while expanding the range of accessible glycans. The instrument drastically shorten and generalizes the synthesis of carbohydrates for use in biomedical and material science

VaporSPOT : parallel synthesis of oligosaccharides on membranes

Automated chemical synthesis has revolutionized synthetic access to biopolymers in terms of simplicity and speed. While automated oligosaccharide synthesis has become faster and more versatile, the parallel synthesis of oligosaccharides is not yet possible. Here, a chemical vapor glycosylation strategy (VaporSPOT) is described that enables the simultaneous synthesis of oligosaccharides on a cellulose membrane solid support. Different linkers allow for flexible and straightforward cleavage, purification, and characterization of the target oligosaccharides. This method is the basis for the development of parallel automated glycan synthesis platforms

The Influence of the Electron Density in Acyl Protecting Groups on the Selectivity of Galactose Formation

The stereoselective formation of 1,2-cis-glycosidic bonds is a major bottleneck in the synthesis of carbohydrates. We here investigate how the electron density in acyl protecting groups influences the stereoselectivity by fine-tuning the efficiency of remote participation. Electron-rich C4-pivaloylated galactose building blocks show an unprecedented α-selectivity. The trifluoroacetylated counterpart with electron-withdrawing groups, on the other hand, exhibits a lower selectivity. Cryogenic infrared spectroscopy in helium nanodroplets and density functional theory calculations revealed the existence of dioxolenium-type intermediates for this reaction, which suggests that remote participation of the pivaloyl protecting group is the origin of the high α-selectivity of the pivaloylated building blocks. According to these findings, an α-selective galactose building block for glycosynthesis is developed based on rational considerations and is subsequently employed in automated glycan assembly exhibiting complete stereoselectivity. Based on the obtained selectivities in the glycosylation reactions and the results from infrared spectroscopy and density functional theory, we suggest a mechanism by which these reactions could proceed

The Influence of the Electron Density in Acyl Protecting Groups on the Selectivity of Galactose Formation

The stereoselective formation of 1,2-cis-glycosidic bonds is a major bottleneck in the synthesis of carbohydrates. We here investigate how the electron density in acyl protecting groups influences the stereoselectivity by fine-tuning the efficiency of remote participation. Electron-rich C4-pivaloylated galactose building blocks show an unprecedented α-selectivity. The trifluoroacetylated counterpart with electron-withdrawing groups, on the other hand, exhibits a lower selectivity. Cryogenic infrared spectroscopy in helium nanodroplets and density functional theory calculations revealed the existence of dioxolenium-type intermediates for this reaction, which suggests that remote participation of the pivaloyl protecting group is the origin of the high α-selectivity of the pivaloylated building blocks. According to these findings, an α-selective galactose building block for glycosynthesis is developed based on rational considerations and is subsequently employed in automated glycan assembly exhibiting complete stereoselectivity. Based on the obtained selectivities in the glycosylation reactions and the results from infrared spectroscopy and density functional theory, we suggest a mechanism by which these reactions could proceed

Thiol-Mediated Uptake of a Cysteine-Containing Nanobody for Anticancer Drug Delivery

The identification of tumor-specific biomarkers is one of the bottlenecks in the development of cancer therapies. Previous work revealed altered surface levels of reduced/oxidized cysteines in many cancers due to overexpression of redox-controlling proteins such as protein disulfide isomerases on the cell surface. Alterations in surface thiols can promote cell adhesion and metastasis, making thiols attractive targets for treatment. Few tools are available to study surface thiols on cancer cells and exploit them for theranostics. Here, we describe a nanobody (CB2) that specifically recognizes B cell lymphoma and breast cancer in a thiol-dependent manner. CB2 binding strictly requires the presence of a nonconserved cysteine in the antigen-binding region and correlates with elevated surface levels of free thiols on B cell lymphoma compared to healthy lymphocytes. Nanobody CB2 can induce complement-dependent cytotoxicity against lymphoma cells when functionalized with synthetic rhamnose trimers. Lymphoma cells internalize CB2 via thiol-mediated endocytosis which can be exploited to deliver cytotoxic agents. CB2 internalization combined with functionalization forms the basis for a wide range of diagnostic and therapeutic applications, rendering thiol-reactive nanobodies promising tools for targeting cancer

VaporSPOT: Parallel Synthesis of Oligosaccharides on Membranes

Automated chemical synthesis has revolutionized synthetic access to biopolymers in terms of simplicity and speed. While automated oligosaccharide synthesis has become faster and more versatile, the parallel synthesis of oligosaccharides is not yet possible. Here, a chemical vapor glycosylation strategy (VaporSPOT) is described that enables the simultaneous synthesis of oligosaccharides on a cellulose membrane solid support. Different linkers allow for flexible and straightforward cleavage, purification, and characterization of the target oligosaccharides. This method is the basis for the development of parallel automated glycan synthesis platforms

Critical Reynolds number for nonlinear flow through rough-walled fractures: The role of shear processes

This paper experimentally investigates the role of shear processes on the variation of critical Reynolds number and nonlinear flow through rough-walled rock fractures. A quantitative criterion was developed to quantify the onset of nonlinear flow by comprehensive combination of Forchheimer's law and Reynolds number. At each shear displacement, several high-precision water flow tests were carried out with different hydraulic gradients then the critical Reynolds number was determined based on the developed criterion. The results show that (i) the Forchheimer's law was fitted very well to experimental results of nonlinear fluid flow through rough-walled fractures, (ii) the coefficients of viscous and inertial pressure drops experience 4 and 7 orders of magnitude reduction during shear displacement, respectively, and (iii) the critical Reynolds number varies from 0.001 to 25 and experiences 4 orders of magnitude enlargement by increasing shear displacement from 0 to 20 mm. These findings may prove useful in proper understanding of fluid flow through rock fractures, or inclusions in computational studies of large-scale nonlinear flow in fractured rocks