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SOLVENT EFFECT ON cDNA-EXPRESSED HUMAN SULFOTRANSFERASE (SULT) ACTIVITIES IN VITRO

Bennett Ma, Magang Shou and Michael L. Schrag
Drug Metabolism and Disposition November 2003, 31 (11) 1300-1305; DOI: https://doi.org/10.1124/dmd.31.11.1300
Bennett Ma
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Magang Shou
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Michael L. Schrag
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Abstract

Sulfation is an important reaction in the biotransformation of steroid hormones, neurotransmitters, drugs, and other xenobiotics, yet little is known about the effects of organic solvents on sulfotransferase (SULT) activities in vitro. Initial experiments found that surprisingly low levels of solvent had dramatic effects on sulfotransferase activity. Consequently, we evaluated the effects of five commonly used solvents (methanol, ethanol, acetonitrile, dimethyl sulfoxide, and dimethyl formamide) on activities of cDNA-expressed sulfotransferase isozymes 1A1 (4-nitrophenol sulfation), 1A3 (dopamine sulfation), 1E1 (ethynylestradiol sulfation), and 2A1 (dehydroepiandrosterone sulfation). In addition, 1-hydroxypyrene was used as a general fluorescent probe for all four sulfotransferase isoforms examined. When substrates were present at their respective isoform-specific Km values, methanol and ethanol (0.4%, v/v) generally had less effect than acetonitrile, dimethyl sulfoxide, and dimethyl formamide on sulfotransferase activities. Acetonitrile, a commonly used solvent in cytochrome P450 studies, inhibited SULT1A1 activities (∼40%) at 0.4% (v/v), but activated SULT1E1-mediated 1-hydroxypyrene sulfation ∼2.6-fold. Assuming a two-site kinetic model, studies revealed that solvent affected Vmax1,Vmax2, and the Ki value of 1-hydroxypyrene sulfation mediated by SULT1E1. In contrast, the Km value was not affected, suggesting that solvent may potentially alter binding interactions of the second substrate molecule, but not the first. Additional experiments with expressed SULT1A1, supplemented with control protein, revealed that the inhibitory effect of solvent (0.4%, v/v) was reduced to <15% for all solvents examined. Thus, it is recommended that ethanol is used as the preferred solvent vehicle and that incubations with expressed enzyme contain >12 μg/ml total protein.

Footnotes

  • ↵1 Abbreviations used are: SULT, sulfotransferase; PAPS, 3′-phosphoadenosine-5′-phosphosulfate; DHEA, dehydroepiandrosterone; HPLC, high-performance liquid chromatography.

    • Received May 5, 2003.
    • Accepted August 21, 2003.
  • The American Society for Pharmacology and Experimental Therapeutics
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Drug Metabolism and Disposition: 31 (11)
Drug Metabolism and Disposition
Vol. 31, Issue 11
1 Nov 2003
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OtherShort Communication

SOLVENT EFFECT ON cDNA-EXPRESSED HUMAN SULFOTRANSFERASE (SULT) ACTIVITIES IN VITRO

Bennett Ma, Magang Shou and Michael L. Schrag
Drug Metabolism and Disposition November 1, 2003, 31 (11) 1300-1305; DOI: https://doi.org/10.1124/dmd.31.11.1300

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OtherShort Communication

SOLVENT EFFECT ON cDNA-EXPRESSED HUMAN SULFOTRANSFERASE (SULT) ACTIVITIES IN VITRO

Bennett Ma, Magang Shou and Michael L. Schrag
Drug Metabolism and Disposition November 1, 2003, 31 (11) 1300-1305; DOI: https://doi.org/10.1124/dmd.31.11.1300
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