High Throughput Screening Tools to Probe and Inhibit Core Fucosylation

We report a chemo-enzymatic transglycosylation synthesis of the 4-methylumbelliferyl glycoside of a complex-type oligosaccharide substrate for core fucosylation. We demonstrate the use of the glycoconjugate in a newly developed enzyme assay to probe the activity and inhibition of fucosyltransferase VIII, which catalyzes the core fucosylation of N-glycans found on eukaryotic glycoproteins. In our assay, the fucosyltransferase VIII reaction is coupled to a specific glycosidase enzyme, allowing to distinguish an unmodified 4-methylumbelliferyl oligosaccharide probe from the fucosylated probe. Our results demonstrate that the assay is very sensitive and specific to the detection of enzymatic activity, while also enabling the identification of potential fucosyltransferase VIII inhibitors

Enzymatic Synthesis of a Fluorogenic Reporter Substrate and a High-Throughput Assay for Fucosyltransferase VIII Provide a Toolkit to Probe and Inhibit Core Fucosylation

We report a straight-forward enzymatic synthesis of the 4-methylumbelliferyl glycoside of a complex-type oligosaccharide substrate for core-fucosylation. We demonstrate the use of this synthetic glycoconjugate in a newly developed enzyme assay to probe the activity and inhibition of fucosyltransferase VIII, which catalyzes the core fucosylation of N-glycans on eukaryotic glycoproteins. In this fucosyltransferase assay, we use the fluorogenic probe and a specific glycosidase in a sequential coupled enzyme reaction to distinguish an unmodified 4-methylumbelliferyl oligosaccharide probe from a fucosylated probe. Our findings show that this strategy is very sensitive and very specific in its detection of enzyme activity and can even be used for analyzing impure tissue lysate samples

The Biology of Vasopressin

Vasopressins are evolutionarily conserved peptide hormones. Mammalian vasopressin functions systemically as an antidiuretic and regulator of blood and cardiac flow essential for adapting to terrestrial environments. Moreover, vasopressin acts centrally as a neurohormone involved in social and parental behavior and stress response. Vasopressin synthesis in several cell types, storage in intracellular vesicles, and release in response to physiological stimuli are highly regulated and mediated by three distinct G protein coupled receptors. Other receptors may bind or cross-bind vasopressin. Vasopressin is regulated spatially and temporally through transcriptional and post-transcriptional mechanisms, sex, tissue, and cell-specific receptor expression. Anomalies of vasopressin signaling have been observed in polycystic kidney disease, chronic heart failure, and neuropsychiatric conditions. Growing knowledge of the central biological roles of vasopressin has enabled pharmacological advances to treat these conditions by targeting defective systemic or central pathways utilizing specific agonists and antagonists