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Abstract

3'-Phosphoadenosine-5'-phosphosulfate: photoaffinity ligand for sulfotransferase enzymes.

D M Otterness, S P Powers, L J Miller and R M Weinshilboum
Molecular Pharmacology January 1991, 39 (1) 34-41;
D M Otterness
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S P Powers
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L J Miller
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R M Weinshilboum
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Abstract

Sulfation is an important pathway in the biotransformation of many drugs, xenobiotic compounds, neurotransmitters, and hormones. The sulfate donor for these reactions is 3'-phosphoadenosine-5'-phosphosulfate (PAPS). We set out to determine whether PAPS might serve as a photoaffinity ligand for sulfotransferase enzymes. UV irradiation of [35S]PAPS with partially purified human liver thermostable (TS) phenol sulfotransferase (PST) radioactively labeled a protein with a molecular mass of 35 kDa, as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Photoaffinity labeling of TS PST with [35S] PAPS did not require the presence of a phenolic substrate but rather was inhibited by p-nitrophenol, a sulfate acceptor substrate for TS PST. Inhibitors of TS PST enzymatic activity, including 3'-phosphoadenosine-5'-phosphate, ATP, ADP, and 2,6-dichloro-4-nitrophenol, also inhibited photoaffinity labeling of the 35-kDa protein with [35S]PAPS, in a concentration-dependent fashion, with IC50 values of 14 microM, 2.1 mM, 7.7 mM, and 91 microM, respectively. The 35-kDa protein that was radioactively labeled by [35S]PAPS in the presence of UV light coeluted with TS PST enzymatic activity during gel filtration high performance liquid chromatography. [35S]PAPS was then used to photoaffinity label another sulfotransferase enzyme, the thermolabile (TL) form of PST partially purified from human liver. Therefore, [35S]PAPS appears to be a photoaffinity ligand that could be used to study a variety of PAPS-dependent sulfotransferases. Photoaffinity labeling of TS and TL PST, as well as other PAPS-dependent sulfotransferases, should enhance our ability to purify this important group of enzymes and to determine amino acid sequences at or near their active sites.

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Molecular Pharmacology
Vol. 39, Issue 1
1 Jan 1991
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Abstract

3'-Phosphoadenosine-5'-phosphosulfate: photoaffinity ligand for sulfotransferase enzymes.

D M Otterness, S P Powers, L J Miller and R M Weinshilboum
Molecular Pharmacology January 1, 1991, 39 (1) 34-41;

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Abstract

3'-Phosphoadenosine-5'-phosphosulfate: photoaffinity ligand for sulfotransferase enzymes.

D M Otterness, S P Powers, L J Miller and R M Weinshilboum
Molecular Pharmacology January 1, 1991, 39 (1) 34-41;
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