title: An HPLC-ECD procedure for measuring total phenolsulfotransferase (PST) activity in human liver, platelets and blood.
abstract: The phenolsulfotransferase (PST) activity in human liver, platelets and blood was measured under saturating concentrations of the conjugating agent, 3'-phosphoadenosine-5'-phosphosulfate (PAPS). Conventional PST assays employ PAP35S at suboptimal concentrations. In addition, the sulfate conjugate formed, namely N-acetyldopamine-sulfate (NADA-sulfate) was quantified directly by high-pressure liquid chromatography cum electrochemical detection (HPLC-ECD). NADA, the biogenic amine acceptor used in this study appeared from kinetic data to be a substrate of both the P and M forms of PST when used in micromolar concentration. Two apparent Km values of 4.2 mumol/l and 22.6 mumol/l were observed. In contrast, only one apparent Km value was evident when the assay was carried out in the presence of 2,6-dichloro-4-nitrophenol (DCNP), a selective inhibitor of the P form of PST or after heat treatment under specified conditions which inactivates the M form of PST. Thus measurement of PST activity with NADA as the acceptor substrate permits the determination of total PST activity and a parallel assay with the inclusion of DCNP would distinguish the two variants of PST, both of which appear to be present in all human tissues.
authors.affiliation: Department of Biochemistry, National University of Singapore.
authors.forename: B Y
authors.initials: BY
authors.lastname: Khoo
authors.forename: K H
authors.initials: KH
authors.lastname: Sit
authors.forename: K P
authors.initials: KP
authors.lastname: Wong
mesh_terms: D015239:Arylsulfotransferase / Q000032:analysis* / Q000097:blood / Q000378:metabolism; D001792:Blood Platelets / Q000201:enzymology*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004298:Dopamine / Q000031:analogs & derivatives / Q000032:analysis / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D010724:Phosphoadenosine Phosphosulfate / Q000378:metabolism; D013462:Sulfur Radioisotopes
chemical_list: D013462:Sulfur Radioisotopes; C068801:N-acetyldopamine-sulfate; D010724:Phosphoadenosine Phosphosulfate; D015239:Arylsulfotransferase; D004298:Dopamine