dynamics simulations demonstrated that UGT94D1-F119I/D188P exhibited a shorter distance between the nucleophilic oxygen (OH6) of the substrate Reb D and uridine diphosphate glucose, along with an increased Ophosphate-C1-Oacceptor angle, thus improving the catalytic activity of the enzyme. Therefor...
The ent-kaurenoic acid biosynthesis pathway: In this pathway, the enzyme ent-copalyl diphosphate synthase (CDPS) first synthesises geranylgeranyl pyrophosphate (GGPP) from ent-copalyl pyrophosphate. Four enzymes control this pathway - CDPS, kaurene oxidase (KO), kaurene synthase (KS) and kaurenoic...
Frozen cells were resuspended in protein buffer (20 mM HEPES pH 8.0, 100 mM NaCl, 1 mM EDTA) supplemented with 10 µM Leupeptin, 1 mM o-Phenanthroline, 0.1 mM PMSF, 10 µM E-64 and 1 mM Pepstatin A. After sonication, cell debris was separated from membranes by...
They found that the UDP-dependent glycosyltransferases (UGTs) UGT76G1 is an enzyme with poor specificity and they engineer it to increase the accumulation of Reb D or Reb M, and reduce the synthesis of the unwanted side-products16. However, the rebaudiosides pro- duction was still low. ...
However, the enzyme was entirely inactivated after 12 h of processing at 50 °C. To enhance its applicability in industrial production, further improvements in its thermal stability are necessary. In this study, we employed the Pythia program [27] and in vitro enzymatic activity assays to ...
Modeled structure-based computational redesign of a glycosyltransferase for the synthesis of rebaudioside D from rebaudioside AComputational enzyme designEnzyme engineeringProtein structure predictionMolecular dynamics simulationGlycosyltransferaseSteviol glycosideEngineering of enzymes without crystal structures and ...
(Svglys/RebA, 5–50 mg/mL), enzyme activity (5–25 U/g Svglys/RebA), temperature (50–70 °C), reaction time (1–6 h) and pH (5–7). The resultant design consisted of 16 experiments to determine the optimal enzyme to proceed with the optimization. Once the optimum enzyme was ...