Reaktion #612970

ord-b708590dea1244efa2e9bf3b0eedc99c

Reaktionsgleichung

CC1OC(OP(=O)(O)OP(=O)(O)OCC2OC(n3cnc4c(=O)nc(N)[nH]c43)C(O)C2O)C(O)C(O)C1O
GDP-fucose
CC(=O)C(=O)[O-]
pyruvate
C[C@H](O)[C@@H](O)[C@@H](O)[C@H](O)C=O
fucose
C=C(OP(=O)(O)O)C(=O)O
Phosphoenolpyruvic acid
Nc1ncnc2c1ncn2[C@@H]1O[C@H](COP(=O)(O)OP(=O)(O)OP(=O)(O)O)[C@@H](O)[C@H]1O
ATP
Nc1nc2c(ncn2[C@@H]2O[C@H](COP(=O)(O)OP(=O)(O)OP(=O)(O)O)[C@@H](O)[C@H]2O)c(=O)[nH]1
GTP
[Cl-].[Cl-].[Mg+2]
MgCl2
Cl.NC(CO)(CO)CO
Tris-HCl
→
CC(=O)N[C@@H]1[C@@H](O[C@@H]2O[C@H](CO)[C@H](O)[C@H](O)[C@H]2O[C@@H]2O[C@@H](C)[C@@H](O)[C@@H](O)[C@@H]2O)[C@H](O)[C@@H](CO)O[C@H]1O
Globo H
Ausbeute 94.0%

Edukte

CC1OC(OP(=O)(O)OP(=O)(O)OCC2OC(n3cnc4c(=O)nc(N)[nH]c43)C(O)C2O)C(O)C(O)C1O
GDP-fucose
CC(=O)C(=O)[O-]
pyruvate
C[C@H](O)[C@@H](O)[C@@H](O)[C@H](O)C=O
fucose
C=C(OP(=O)(O)O)C(=O)O
Phosphoenolpyruvic acid
Nc1ncnc2c1ncn2[C@@H]1O[C@H](COP(=O)(O)OP(=O)(O)OP(=O)(O)O)[C@@H](O)[C@H]1O
ATP
Nc1nc2c(ncn2[C@@H]2O[C@H](COP(=O)(O)OP(=O)(O)OP(=O)(O)O)[C@@H](O)[C@H]2O)c(=O)[nH]1
GTP
[Cl-].[Cl-].[Mg+2]
MgCl2
Cl.NC(CO)(CO)CO
Tris-HCl

Reagenzien

Keine

Reaktionsbedingungen

Detaillierte Bedingungen
See reaction.notes.procedure_details.

Aufarbeitung

  1. 1
    Sonstigewas placed into an incubator at 16˜50° C. with gentle shaking
  2. 2
    workup.ADDITIONMore enzymes are added if the reaction
  3. 3
    SonstigeThe Globo H product was purified by 18C reverse phase column in 94% yield

Vorschrift

5 mmol Gb5 with linker, 5 mmol fucose, 12 mmol Phosphoenolpyruvic acid (PEP), 0.25 mmol ATP, 0.25 mmol GTP with 10 mM MgCl2 were added into 100 mM Tris-HCl buffer (pH 7.5). The reaction was initiated by addition suitable amount of α-1,2-fucosyltransferase, L-fucokinase/GDP-fucose pyrophosphorylase (FKP), pyruvate kinase (PK) and pyrophosphatase (PPA). The flask was placed into an incubator at 16˜50° C. with gentle shaking. The reaction was monitored by TLC. More enzymes are added if the reaction stops. The reaction is stopped when no more starting material is observed by TLC. The Globo H product was purified by 18C reverse phase column in 94% yield.

Quelle

DOI: 10.6084/m9.figshare.5104873.v1Patent: US09340812B2uspto-grants-2016_05