Search

In this study, we developed an efficient fluorous tagging strategy for site-selective enzymatic fucosylation/sialylation in which appending a readily removable auxiliary tag (SF17) to the reducing ...

Bacteroides fragilis α1,3/4-fucosyltransferase (Bf13FT) was expressed in Escherichia coli and characterized as an α1,3/4-fucosyltransferase that can be used as a versatile catalyst for the synthesi...

In this study, we focused on the synthetic application of α1,3/4-fucosyltransferase obtained from Helicobacter pylori DSM 6709 (FucTIII) on l-fucose-containing glycans. By combining FucTIII with the sequential one-pot enzymatic system of human milk oligosaccharide (HMO) production, various fucosylated HMOs, such as lacto-N-fucopentose V (LNFP V), LNFP VI, lacto-N-difucohexaose II (LNDFH II), and lacto-N-neodifucohexaose II (LNnDFH II), were synthesized. Moreover, l-fucose-containing glycan synthesis of Lewis antigens, such as Lewis x, Lewis y, Lewis a, Lewis b, sialyl Lewis x, sialyl Lewis a, and their derivatives, was achieved. Enzyme kinetics proved that the catalytic efficiency (kcat/Km) of FucTIII on type-2 N-acetyl lactosamine (LacNAc) was 39 times higher than that on type-1 LacNAc. Furthermore, enzyme kinetics revealed that additional GlcNAc on the nonreducing end of the acceptors can enhance the catalytic efficiency of FucTIII on the glycan acceptors. Investigations of bacterial FucTs on substrate ...

In cancers, increased fucosylation (attachment of fucose sugar residues) on cell-surface glycans, resulting from the abnormal upregulation of the expression of specific fucosyltransferase enzymes (FUTs), is one of the most important types of glycan modifications associated with malignancy. Fucosylated glycans on cell surfaces are involved in a multitude of cellular interactions and signal regulation in normal biological processes, as well as in disease. For example, sialyl LewisX is a fucosylated cell-surface glycan that is abnormally abundant in some cancers where it has been implicated in facilitating metastasis, allowing circulating tumor cells to bind to the epithelial tissue within blood vessels and invade into secondary sites by taking advantage of glycan-mediated interactions. To identify inhibitors of FUT enzymes as potential cancer therapeutics, we have developed a novel high-throughput assay that makes use of a fluorogenically labeled oligosaccharide as a probe of fucosylation. This probe, which consists of a 4-methylumbelliferyl glycoside, is recognized and hydrolyzed by specific glycoside hydrolase enzymes to release fluorescent 4-methylumbelliferone, yet when the probe is fucosylated prior to treatment with the glycoside hydrolases, hydrolysis does not occur and no fluorescent signal is produced. We have demonstrated that this assay can be used to measure the inhibition of FUT enzymes by small molecules, because blocking fucosylation will allow glycosidase-catalyzed hydrolysis of the labeled oligosaccharide to produce a fluorescent signal. Employing this assay, we have screened a focused library of small molecules for inhibitors of a human FUT enzyme involved in the synthesis of sialyl LewisX and demonstrated that our approach can be used to identify potent FUT inhibitors from compound libraries in microtiter plate format.

Control of morphology in the synthesis of Rh nanocrystals can be used to precisely tailor the electronic surface structure; this in turn directly influences their performance in catalysis applications. Many works bring attention to the development of Rh nanostructures with low-index surfaces, but limited effort has been placed on the study of high-index and surface-defect-enriched nanocrystals as they are not favored by thermodynamics due to the involvement of high-energy surfaces and increased surface-to-volume ratios. In this work, we demonstrate an aqueous synthesis of concave Rh nanotetrahedra (CTDs) serving as efficient catalysts for energy conversion reactions. CTDs are surface-defect-rich structures that form through a slow growth rate and follow the four-step model of metallic nanoparticle growth. By tuning the surfactant concentration, the morphology of Rh CTDs evolved into highly excavated nanotetrahedra (HETDs) and twinned nanoparticles (TWs). Unlike the CTD surfaces with abundant adatoms and v...

Magnetic nanoparticles (MNPs) are attractive materials that serve as a support for enzyme immobilization and facilitate separations by applying an external magnetic field; this could facilitate the recycling of enzymes and broaden their applications in organic synthesis. Herein, we report the methods for the immobilization of water-soluble and membrane-bound enzymes, and the activity difference between free and immobilized enzymes is discussed. Sialyltransferase (PmST1, from Pasteurella multocida ) and cytidine monophosphate (CMP)-sialic acid synthetase (CSS, from Neisseria meningitides ) were chosen as water-soluble enzymes and expressed using an intein expression system. The enzymes were site-specifically and covalently immobilized on PEGylated-N-terminal cysteine MNPs through native chemical ligation (NCL). Increasing the length of the PEG linker between the enzyme and the MNP surface increased the activity of the immobilized enzymes relative to the free parent enzymes. In addition, the use of a fluorescent acceptor tag for PmST1 affected enzyme kinetics. In contrast, sialyltransferase from Neisseria gonorrheae (NgST, a membrane-bound enzyme) was modified with a biotin-labeled cysteine at the C-terminus using NCL, and the enzyme was then assembled on streptavidin-functionalized MNPs. Using a streptavidin-biotin interaction, it was possible to immobilize NgST on a solid support under mild ligation conditions, which prevented the enzyme from high-temperature decomposition and provided an approximately 2-fold increase in activity compared to other immobilization methods on MNPs. Finally, the ganglioside GM3-derivative (sialyl-lactose derivative) was synthesized in a one-pot system by combining the use of immobilized PmST1 and CSS. The enzymes retained 50% activity after being reused ten times. Furthermore, the results obtained using the one-pot two-immobilized-enzyme system demonstrated that it can be applied to large-scale reactions with acceptable yields and purity. These features make enzyme-immobilized MNPs applicable to organic synthesis.