Title: Study of enzyme kinetics of phenol sulfotransferase by electrophoretically mediated microanalysis
Authors: Nováková, Sona ×
Van Dyck, Sigrid
Glatz, Zdenek
Van Schepdael, Ann
Hoogmartens, Jos #
Issue Date: Apr-2004
Series Title: Journal of Chromatography A vol:1032 issue:1-2 pages:319-26
Abstract: Electrophoretically mediated microanalysis (EMMA) was applied for the study of the kinetic parameters of the enzymatic reaction of phenol sulfotransferase SULT1A1 isoenzyme with 4-nitrophenol as a substrate. The SULT1A1 activity was determined by the quantitation of the product, 4-nitrophenyl sulfate, at 274 nm by using different injection and separation steps. This new approach solved the problem of the presence of the very strong inhibitor, adenosine 3',5'-bisphosphate (PAP), in the co-substrate solution (adenosine 3'-phosphate 5'-phosphosulfate, PAPS) which is unstable at room temperature. The inhibitor PAP was electrophoretically separated from the co-substrate PAPS before the injection of enzyme and substrate inside the capillary (and thus before their in-capillary encountering). With the developed in-capillary SULT1A1 activity assay an average Michaelis constant (Km) for 4-nitrophenol was calculated to be 0.84 microM, a value which is consistent with a previously reported value. Strong substrate inhibition (above a 4-nitrophenol concentration of 2.5 microM) was observed, and this is also in accordance with literature values.
ISSN: 0021-9673
Publication status: published
KU Leuven publication type: IT
Appears in Collections:Pharmaceutical Analysis
× corresponding author
# (joint) last author

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