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. 1994 Sep 1;302(Pt 2):497–502. doi: 10.1042/bj3020497

Functional characterization of two human sulphotransferase cDNAs that encode monoamine- and phenol-sulphating forms of phenol sulphotransferase: substrate kinetics, thermal-stability and inhibitor-sensitivity studies.

M E Veronese 1, W Burgess 1, X Zhu 1, M E McManus 1
PMCID: PMC1137255  PMID: 8093002

Abstract

The present paper describes the functional characterization of two human aryl sulphotransferase (HAST) cDNAs, HAST1 and HAST3, previously isolated by us from liver and brain, respectively [Zhu, Veronese, Sansom, and McManus (1993) Biochem. Biophys. Res. Commun. 192, 671-676; Zhu, Veronese, Bernard, Sansom and McManus (1993) Biochem. Biophys. Res. Commun. 195, 120-127]. These appear to encode the two major forms of phenol sulphotransferase (PST) characterized in a number of human tissue cytosols, these being the phenolsulphating (P-PST) and monoamine-sulphating (M-PST) forms of phenol sulphotransferase. HAST1 and HAST3 cDNAs were functionally expressed in COS-7 cells and kinetically characterized using the model substrates for P-PST and M-PST, p-nitrophenol and dopamine (3,4-dihydroxyphenethylamine) respectively. COS-expressed HAST1 was shown to be enzymatically active in sulphating p-nitrophenol with high affinity (Km 0.6 microM), whereas dopamine was the preferred substrate for HAST3 (Km 9.7 microM). HAST1 could also sulphate dopamine, as could HAST3 sulphate p-nitrophenol, but the Km for these reactions were at least two orders of magnitude greater than for the preferred substrates. COS-expressed HAST1 and HAST3 displayed inhibition profiles with the ST inhibitor 2,6-dichloro-4-nitrophenol (DCNP), identical with human liver cytosolic P-PST and M-PST activities respectively. Thermal-stability studies with the expressed enzymes showed that HAST1 was considerably more thermostable (TS) than HAST3, which is consistent with P-PST being termed the TS PST and M-PST being termed the thermolabile (TL) PST. Western immunoblot analyses of the expressed PST proteins using an antibody generated to a bacterially expressed rat liver aryl/phenol ST showed that HAST1 and HAST3 migrated as single proteins with different electrophoretic mobilities (32 versus 34 kDa). This is consistent with the differences in electrophoretic mobilities observed for P-PST and M-PST in a variety of tissues reported by other workers. This report on the functional characterization of P-PST and M-PST cDNAs provides important information on the structural as well as functional relationships of human PSTs, which sulphate a vast array of exogenous and endogenous compounds.

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Selected References

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  1. Campbell N. R., Van Loon J. A., Weinshilboum R. M. Human liver phenol sulfotransferase: assay conditions, biochemical properties and partial purification of isozymes of the thermostable form. Biochem Pharmacol. 1987 May 1;36(9):1435–1446. doi: 10.1016/0006-2952(87)90108-0. [DOI] [PubMed] [Google Scholar]
  2. Comer K. A., Falany J. L., Falany C. N. Cloning and expression of human liver dehydroepiandrosterone sulphotransferase. Biochem J. 1993 Jan 1;289(Pt 1):233–240. doi: 10.1042/bj2890233. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Cruickshank D., Sansom L. N., Veronese M. E., Mojarrabi B., McManus M. E., Zhu X. cDNA expression studies of rat liver aryl sulphotransferase. Biochem Biophys Res Commun. 1993 Feb 26;191(1):295–301. doi: 10.1006/bbrc.1993.1216. [DOI] [PubMed] [Google Scholar]
  4. Falany C. N. Molecular enzymology of human liver cytosolic sulfotransferases. Trends Pharmacol Sci. 1991 Jul;12(7):255–259. doi: 10.1016/0165-6147(91)90566-b. [DOI] [PubMed] [Google Scholar]
  5. Falany C. N., Vazquez M. E., Heroux J. A., Roth J. A. Purification and characterization of human liver phenol-sulfating phenol sulfotransferase. Arch Biochem Biophys. 1990 May 1;278(2):312–318. doi: 10.1016/0003-9861(90)90265-z. [DOI] [PubMed] [Google Scholar]
  6. Foldes A., Meek J. L. Rat brain phenolsulfotransferase: partial purification and some properties. Biochim Biophys Acta. 1973 Dec 19;327(2):365–374. doi: 10.1016/0005-2744(73)90419-1. [DOI] [PubMed] [Google Scholar]
  7. Heroux J. A., Falany C. N., Roth J. A. Immunological characterization of human phenol sulfotransferase. Mol Pharmacol. 1989 Jul;36(1):29–33. [PubMed] [Google Scholar]
  8. McManus M. E., Hall P. D., Stupans I., Brennan J., Burgess W., Robson R., Birkett D. J. Immunohistochemical localization and quantitation of NADPH-cytochrome P-450 reductase in human liver. Mol Pharmacol. 1987 Aug;32(1):189–194. [PubMed] [Google Scholar]
  9. Otterness D. M., Wieben E. D., Wood T. C., Watson W. G., Madden B. J., McCormick D. J., Weinshilboum R. M. Human liver dehydroepiandrosterone sulfotransferase: molecular cloning and expression of cDNA. Mol Pharmacol. 1992 May;41(5):865–872. [PubMed] [Google Scholar]
  10. Reiter C., Mwaluko G., Dunnette J., Van Loon J., Weinshilboum R. Thermolabile and thermostable human platelet phenol sulfotransferase. Substrate specificity and physical separation. Naunyn Schmiedebergs Arch Pharmacol. 1983 Sep;324(2):140–147. doi: 10.1007/BF00497020. [DOI] [PubMed] [Google Scholar]
  11. Sundaram R. S., Szumlanski C., Otterness D., van Loon J. A., Weinshilboum R. M. Human intestinal phenol sulfotransferase: assay conditions, activity levels and partial purification of the thermolabile form. Drug Metab Dispos. 1989 May-Jun;17(3):255–264. [PubMed] [Google Scholar]
  12. Weinshilboum R. Sulfotransferase pharmacogenetics. Pharmacol Ther. 1990;45(1):93–107. doi: 10.1016/0163-7258(90)90010-y. [DOI] [PubMed] [Google Scholar]
  13. Whittemore R. M., Pearce L. B., Roth J. A. Purification and kinetic characterization of a dopamine-sulfating form of phenol sulfotransferase from human brain. Biochemistry. 1985 May 7;24(10):2477–2482. doi: 10.1021/bi00331a013. [DOI] [PubMed] [Google Scholar]
  14. Whittemore R. M., Pearce L. B., Roth J. A. Purification and kinetic characterization of a phenol-sulfating form of phenol sulfotransferase from human brain. Arch Biochem Biophys. 1986 Sep;249(2):464–471. doi: 10.1016/0003-9861(86)90023-8. [DOI] [PubMed] [Google Scholar]
  15. Wilborn T. W., Comer K. A., Dooley T. P., Reardon I. M., Heinrikson R. L., Falany C. N. Sequence analysis and expression of the cDNA for the phenol-sulfating form of human liver phenol sulfotransferase. Mol Pharmacol. 1993 Jan;43(1):70–77. [PubMed] [Google Scholar]
  16. Young W. F., Jr, Okazaki H., Laws E. R., Jr, Weinshilboum R. M. Human brain phenol sulfotransferase: biochemical properties and regional localization. J Neurochem. 1984 Sep;43(3):706–715. doi: 10.1111/j.1471-4159.1984.tb12790.x. [DOI] [PubMed] [Google Scholar]
  17. Zhu X., Veronese M. E., Bernard C. C., Sansom L. N., McManus M. E. Identification of two human brain aryl sulfotransferase cDNAs. Biochem Biophys Res Commun. 1993 Aug 31;195(1):120–127. doi: 10.1006/bbrc.1993.2018. [DOI] [PubMed] [Google Scholar]
  18. Zhu X., Veronese M. E., Sansom L. N., McManus M. E. Molecular characterisation of a human aryl sulfotransferase cDNA. Biochem Biophys Res Commun. 1993 Apr 30;192(2):671–676. doi: 10.1006/bbrc.1993.1467. [DOI] [PubMed] [Google Scholar]

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