Yu-Hsuan Tsai

Facile synthesis of size dependent Ru(II)-carbohydrate dendrimers via click chemistry.

A facile and flexible approach for the preparation of Ru(II) complexes containing different carbohydrates based on the Cu(II)-catalyzed Huisgen-[3+2] cycloaddition is described.

Selective, rapid and optically switchable regulation of protein function in live mammalian cells

The rapid and selective regulation of a target protein within living cells that contain closely related family members is an outstanding challenge. Here we introduce genetically directed bioorthogonal ligand tethering (BOLT) and demonstrate selective inhibition (iBOLT) of protein function. In iBOLT, inhibitor-conjugate/target protein pairs are created where the target protein contains a genetically encoded unnatural amino acid with bioorthogonal reactivity and the inhibitor conjugate contains a complementary bioorthogonal group. iBOLT enables the first rapid and specific inhibition of MEK isozymes, and introducing photoisomerizable linkers in the inhibitor conjugate enables reversible, optical regulation of protein activity (photo-BOLT) in live mammalian cells. We demonstrate that a pan kinase inhibitor conjugate allows selective and rapid inhibition of the lymphocyte specific kinase, indicating the modularity and scalability of BOLT. We anticipate that BOLT will enable the rapid and selective regulation of diverse proteins for which no selective small-molecule ligands exist.

Molecular analysis of Carbohydrate−Antibody interactions : case study using a Bacillus anthracis Tetrasaccharide

The process for selecting potent and effective carbohydrate antigens is not well-established. A combination of synthetic glycan microarray screening, surface plasmon resonance analysis, and saturation transfer difference NMR spectroscopy was used to dissect the antibody-binding surface of a carbohydrate antigen, revealing crucial binding elements with atomic-level detail. This analysis takes the first step toward uncovering the rules for structure-based design of carbohydrate antigens.

Chemical biology of glycosylphosphatidylinositol anchors

Glycosylphosphatidylinositols (GPIs) are complex glycolipids that are covalently linked to the C-terminus of proteins as a posttranslational modification. They anchor the attached protein to the cell membrane and are essential for normal functioning of eukaryotic cells. GPI-anchored proteins are structurally and functionally diverse. Many GPIs have been structurally characterized but comprehension of their biological functions, beyond the simple physical anchoring, remains largely speculative. Work on functional elucidation at a molecular level is still limited. This Review focuses on the roles of GPI unraveled by using synthetic molecules and summarizes the structural diversity of GPIs, as well as their biological and chemical syntheses.

A general and convergent synthesis of diverse glycosylphosphatidylinositol glycolipids

Glycosylphosphatidylinositol (GPI) glycolipids anchor a large number of proteins in the cell membrane of eukaryotic cells. Their conserved pseudopentasaccharide core carries additional phosphoethanolamine, saccharide and lipid substituents. These structural variations are characteristic for a species or a tissue but their functional significance remains largely unknown. Studies that would link a specific function to a structurally unique GPI rely on availability of homogeneous samples of these glycolipids. To address this need we have developed a general synthetic route to GPI glycolipids. Our convergent synthesis starts from common building blocks and relies on a fully orthogonal set of protecting groups that enables the regioselective introduction of phosphodiesters and efficient assembly of the GPI glycans. Here, we report on the development of this synthetic strategy, evaluation of the set of protecting groups with respect to phosphorylation methods, evaluation of the assembly plan for the GPI glycan, optimization of the glycosylation reactions, and the application of this strategy to the total syntheses of four structurally diverse branched GPI glycolipids.

A General Method for Synthesis of GPI Anchors Illustrated by the Total Synthesis of the Low‐Molecular‐Weight Antigen from Toxoplasma gondii

Building blocks: a new, general synthetic strategy, which allows the construction of branched glycosylphosphatidylinositols (GPIs), enables the synthesis of parasitic glycolipid 1 from Toxoplasma gondii. In addition, the structure is further confirmed by recognition of monoclonal antibodies.

Diagnosis of toxoplasmosis using a synthetic glycosylphosphatidylinositol glycan.

Around 2 billion people worldwide are infected with the apicomplexan parasite Toxoplasma gondii which induces a variety of medical conditions. For example, primary infection during pregnancy can result in fetal death or mental retardation of the child. Diagnosis of acute infections in pregnant women is challenging but crucially important as the drugs used to treat T. gondii infections are potentially harmful to the unborn child. Better, faster, more reliable, and cheaper means of diagnosis by using defined antigens for accurate serological tests are highly desirable. Synthetic pathogen-specific glycosylphosphatidylinositol (GPI) glycan antigens are diagnostic markers and have been used to distinguish between toxoplasmosis disease states using human sera.

Synthetic Inositol Phosphoglycans Related to GPI Lack Insulin-Mimetic Activity

Insulin signaling has been suggested, at least in part, to be affected by an insulin-mimetic species of low molecular weight. These inositol phosphoglycans (IPGs) are generated upon growth hormone/cytokine stimulation and control the activity of a multitude of insulin effector enzymes. The minimal structural requirements of IPGs for insulin-mimetic action have been debated. Two types of IPGs were suggested, and the IPG-A type resembles the core glycan of glycosylphosphatidylinositol (GPI)-anchors. In fact, purified GPI-anchors of lower eukaryotic origin have been shown to influence glucose homeostasis. To elucidate active IPGs, a collection of synthetic IPGs designed on the basis of previous reports of activity were tested for their insulin-mimetic activity. In vitro and ex vivo assays in rodent adipose tissue as well as in vivo analyses in mice were employed to test the synthetic IPGs. None of the IPGs we tested mimic insulin actions as determined by PKB/Akt phosphorylation and quantification of glucose transport and lipogenesis. Furthermore, none of the IPGs had any effect in in vivo insulin tolerance assays. In stark contrast to previous claims, we conclude that neither of the compounds tested is insulin-mimetic.

Structural variation of glycolipids from Meiothermus taiwanensis ATCC BAA-400 under different growth temperatures

A major glycolipid, alpha-Galf(1-3)-alpha-Galp(1-6)-beta-GlcpNAcyl(1-2)-alpha-Glcp(1-1)-2-acylalkyldiol, is obtained from Meiothermus taiwanensis. This novel glycolipid is found only when the bacterium grows above 62 degrees C, which is significantly different from those from the same bacteria incubated at 55 degrees C. Terminal galactofuranoside and 1,2-alkyldiol lipids replaced galactopyranoside and glycerol lipids, respectively, under increased growth temperature. This variation is likely necessary for bacteria for keeping the stable outer membrane and surviving under extreme environments.

Investigation of the protective properties of glycosylphosphatidylinositol-based vaccine candidates in a Toxoplasma gondii mouse challenge model

Vaccination against the ubiquitous parasite Toxoplasma gondii would provide the most efficient prevention against toxoplasmosis-related congenital, brain and eye diseases in humans. We investigated the immune response elicited by pathogen-specific glycosylphosphatidylinositol (GPI) glycoconjugates using carbohydrate microarrays in a BALB/c mouse model. We further examined the protective properties of the glycoconjugates in a lethal challenge model using the virulent T. gondii RH strain. Upon immunization, mice raised antibodies that bind to the respective GPIs on carbohydrate microarrays, but were mainly directed against an unspecific GPI epitope including the linker. The observed immune response, though robust, was unable to provide protection in mice when challenged with a lethal dose of viable tachyzoites. We demonstrate that anti-GPI antibodies raised against the here described semi-synthetic glycoconjugates do not confer protective immunity against T. gondii in BALB/c mice.