40591-51-3

Upstream product

• 25546-57-0 dibenzyl (2,3,4,6-tetra-O-acetyl-β-D-glucopyranosyl) phosphate 
• 572-09-8 2,3,4,6-tetra-O-acetyl-α-D-glucopyranosyl bromide 
• 38768-84-2 Dibenzyl(2,3,4,6-tetra-O-benzyl-β-D-glucopyranosyl)phosphat 
• 82300-63-8 Dibenzyl(β-D-glucopyranosyl)phosphat 
• 58526-05-9 2,3,4,6-tetra-O-acetyl-β-D-glucopyranosyl phosphate 
• 64-17-5 ethanol 
• 74808-10-9 O-(2,3,4,6-Tetra-O-acetyl-α-D-glucopyranosyl)trichloroacetimidate 
• 4768-77-8 exo-3,4,6-tri-O-acetyl-1,2-O-(tert-butyl orthoacetyl)-α-D-glucopyranose 
• 492-62-6 alpha-D-glucopyranose 
• 2520-52-7 α-D-galactopyranosyl phosphate 
• 69-79-4 D-maltose 
• 3068-32-4 1-bromo-1-deoxy-2,3,4,6-tetra-O-acetyl-a-D-galactopyranoside 
• 170428-70-3 3-methoxy-2-pyridyl β-D-galactopyranoside 
• 71-43-2 benzene 

Downstream Products

• 14034-70-9 β-D-glucopyranosyl 
• 69557-93-3 O-α-D-Glucopyranosyl-(1-3)-α,β-L-rhamnopyranose 
• 492-62-6 alpha-D-glucopyranose 
• 2520-52-7 α-D-galactopyranosyl phosphate 
• 492-61-5 β-D-glucose 
• 50692-66-5 benzyl-2-acetamido-2-deoxy-3-O-(β-D-galactopyranosyl)-α-D-glucopyranoside 
• 50787-09-2 β-Galp-(1->3)-GlcNAc 
• 19046-60-7 α-D-galactopyranosyl phosphate dipotassium salt 
• 133-89-1 UDP-glucose 
• 1242712-29-3 5-(4-methoxyphenyl)-UDP-Gal 
• 991-43-5 guanosine 5'-diphospho-D-mannose 
• 154235-70-8 guanosine 5’-diphospho-β-L-fucose 
• 133-89-1 Uridine diphosphate mannose 
• 14034-70-9 α-D-mannopyranosyl (bis(cyclohexylammonium)phosphate) 

Relevant academic research and scientific papers

β-Glucose-1,6-Bisphosphate stabilizes pathological phophomannomutase2 mutants in vitro and represents a lead compound to develop pharmacological chaperones for the most common disorder of glycosylation, PMM2-CDG

A large number of mutations causing PMM2-CDG, which is the most frequent disorder of glycosylation, destabilize phosphomannomutase2. We looked for a pharmacological chaperone to cure PMM2-CDG, starting from the structure of a natural ligand of phosphomannomutase2, α-glucose-1,6-bisphosphate. The compound, β-glucose-1,6-bisphosphate, was synthesized and characterized via 31P-NMR. β-glucose-1,6-bisphosphate binds its target enzyme in silico. The binding induces a large conformational change that was predicted by the program PELE and validated in vitro by limited proteolysis. The ability of the compound to stabilize wild type phosphomannomutase2, as well as frequently encountered pathogenic mutants, was measured using thermal shift assay. β-glucose-1,6-bisphosphate is relatively resistant to the enzyme that specifically hydrolyses natural esose-bisphosphates.

Toward Automated Enzymatic Glycan Synthesis in a Compartmented Flow Microreactor System

Immobilized microfluidic enzyme reactors (IMER) are of particular interest for automation of enzyme cascade reactions. Within an IMER, substrates are converted by paralleled immobilized enzyme modules and intermediate products are transported for further conversion by subsequent enzyme modules. By optimizing substrate conversion in the spatially separated enzyme modules purification of intermediate products is not necessary, thus shortening process time and increasing space-time yields. The IMER enables the development of efficient enzyme cascades by combining compatible enzymatic reactions in different arrangements under optimal conditions and the possibility of a cost-benefit analysis prior to scale-up. These features are of special interest for automation of enzymatic glycan synthesis. We here demonstrate a compartmented flow microreactor system using six magnetic enzyme beads (MEBs) for the synthesis of the non-sulfated human natural killer cell-1 (HNK-1) glycan epitope. MEBs are assembled to build compartmented enzyme modules, consisting of enzyme cascades for the synthesis of uridine 5′- diphospho-α- d-galactose (UDP-Gal) and uridine 5′-diphospho-α-d-glucuronic acid (UDP-GlcA), the donor substrates for the Leloir glycosyltransferases β4-galactosyltransferase and β3-glucuronosyltransferase, respectively. Glycan synthesis was realized in an automated microreactor system by a cascade of individual enzyme module compartments each performing under optimal conditions. The products were analyzed inline by an MS-system connected to the microreactor. The high synthesis yield of 96% for the non-sulfated HNK-1 glycan epitope indicates the excellent performance of the automated enzyme module cascade. Furthermore, combinations of other MEBs for nucleotide sugars synthesis with MEBs of glycosyltransferases have the potential for a fully automated and programmed glycan synthesis in a compartmented flow microreactor system. (Figure presented.).

A mutant of phosphomannomutase1 retains full enzymatic activity, but is not activated by IMP: Possible implications for the disease PMM2-CDG

The most frequent disorder of glycosylation, PMM2-CDG, is caused by a deficiency of phosphomannomutase activity. In humans two paralogous enzymes exist, both of them require mannose 1,6-bis-phosphate or glucose 1,6-bis-phosphate as activators, but only phospho-mannomutase1 hydrolyzes bis-phosphate hexoses. Mutations in the gene encoding phos-phomannomutase2 are responsible for PMM2-CDG. Although not directly causative of the disease, the role of the paralogous enzyme in the disease should be clarified. Phosphoman-nomutase1 could have a beneficial effect, contributing to mannose 6-phosphate isomerization, or a detrimental effect, hydrolyzing the bis-phosphate hexose activator. A pivotal role in regulating mannose-1phosphate production and ultimately protein glycosylation might be played by inosine monophosphate that enhances the phosphatase activity of phosphoman-nomutase1. In this paper we analyzed human phosphomannomutases by conventional enzymatic assays as well as by novel techniques such as 31P-NMR and thermal shift assay. We characterized a triple mutant of phospomannomutase1 that retains mutase and phosphatase activity, but is unable to bind inosine monophosphate.

Facile enzymatic synthesis of sugar 1-phosphates as substrates for phosphorylases using anomeric kinases

Three sugar 1-phosphates that are donor substrates for phosphorylases were produced at the gram scale from phosphoenolpyruvic acid and the corresponding sugars by the combined action of pyruvate kinase and the corresponding anomeric kinases in good yields. These sugar 1-phosphates were purified through two electrodialysis steps. α-d-Galactose 1-phosphate was finally isolated as crystals of dipotassium salts. α-d-Mannose 1-phosphate and 2-acetamido-2-deoxy-α-d-glucose 1-phosphate were isolated as crystals of bis(cyclohexylammonium) salts.

Efficient chemoenzymatic synthesis of novel galacto-N-biose derivatives and their sialylated forms

Galacto-N-biose (GNB) derivatives were efficiently synthesized from galactose derivatives via a one-pot two-enzyme system containing two promiscuous enzymes from Bifidobacterium infantis: a galactokinase (BiGalK) and a d-galactosyl-β1-3-N-acetyl-d-hexosamine phosphorylase (BiGalHexNAcP). Mono-sialyl and di-sialyl galacto-N-biose derivatives were then prepared using a one-pot two-enzyme system containing a CMP-sialic acid synthetase and an α2-3-sialyltransferase or an α2-6-sialyltransferase.

Engineering the specificity of trehalose phosphorylase as a general strategy for the production of glycosyl phosphates

A two-step process is reported for the anomeric phosphorylation of galactose, using trehalose phosphorylase as biocatalyst. The monosaccharide enters this process as acceptor but can subsequently be released from the donor side, thanks to the non-reducing nature of the disaccharide intermediate. A key development was the creation of an optimized enzyme variant that displays a strict specificity (99%) for β-galactose 1-phosphate as product. This journal is the Partner Organisations 2014.

A highly efficient galactokinase from Bifidobacterium infantis with broad substrate specificity

Galactokinase (GalK), particularly GalK from Escherichia coli, has been widely employed for the synthesis of sugar-1-phosphates. In this study, a GalK from Bifidobacterium infantis ATCC 15697 (BiGalK) was cloned and over-expressed with a yield of over 80 mg/L cell cultures. The kcat/Km value of recombinant BiGalK toward galactose (164 s-1 mM -1) is 296 times higher than that of GalK from E. coli, indicating that BiGalK is much more efficient in the phosphorylation of galactose. The enzyme also exhibits activity toward galacturonic acid, which has never been observed on other wild type GalKs. Further activity assays showed that BiGalK has broad substrate specificity toward both sugars and phosphate donors. These features make BiGalK an attractive candidate for the large scale preparation of galactose-1-phosphate and derivatives.

Substrate specificity of galactokinase from Streptococcus pneumoniae TIGR4 towards galactose, glucose, and their derivatives

Galactokinases (GalKs) have attracted significant research attention for their potential applications in the enzymatic synthesis of unique sugar phosphates. The galactokinase (GalKSpe4) cloned from Streptococcus pneumoniae TIGR4 presents a remarkably broad substrate range including 14 diverse natural and unnatural sugars. TLC and MS studies revealed that GalKSpe4 had relaxed activity towards galactose derivatives with modifications on the C-6, 4- or 2-positions. Additionally, GalKSpe4 can also tolerate glucose while glucose derivatives with modifications on the C-6, 4- or 2-positions were unacceptable. More interestingly, GalKSpe4 can phosphorylate l-mannose in moderate yield (43%), while other l-sugars such as l-Gal cannot be recognized by this enzyme. These results are very significant because there is rarely enzyme reported that can phosphorylate such uncommon substrates as l-mannose.

The role of the active site residues in human galactokinase: Implications for the mechanisms of GHMP kinases

Galactokinase catalyses the phosphorylation of galactose at the expense of ATP. Like other members of the GHMP family of kinases it is postulated to function through an active site base mechanism in which Asp-186 abstracts a proton from galactose. This asparate residue was altered to alanine and to asparagine by site-directed mutagenesis of the corresponding gene. This resulted in variant enzyme with no detectable galactokinase activity. Alteration of Arg-37, which lies adjacent to Asp-186 and is postulated to assist the catalytic base, to lysine resulted in an active enzyme. However, alteration of this residue to glutamate abolished activity. All the variant enzymes, except the arginine to lysine substitution, were structurally unstable (as judged by native gel electrophoresis in the presence of urea) compared to the wild type. This suggests that the lack of activity results from this structural instability, in addition to any direct effects on the catalytic mechanism. Computational estimations of the pKa values of the arginine and aspartate residues, suggest that Arg-37 remains protonated throughout the catalytic cycle whereas Asp-186 has an abnormally high pKa value (7.18). Quantum mechanics/molecular mechanics (QM/MM) calculations suggest that Asp-186 moves closer to the galactose molecule during catalysis. The experimental and theoretical studies presented here argue for a mechanism in which the C 1-OH bond in the sugar is weakened by the presence of Asp-186 thus facilitating nucleophilic attack by the oxygen atom on the γ-phosphorus of ATP.