Expression and Characterization of a Novel Thyroid Hormone-Sulfating Form of Cytosolic Sulfotransferase from Human Liver
- Department of Pharmacology and Toxicology, University of Alabama at Birmingham, Birmingham, Alabama 35294
Abstract
Sulfation is an important conjugation reaction for a wide range of endogenous and exogenous compounds in humans, including steroids, bile acids, catecholamine neurotransmitters and thyroid hormones. The cDNA for a distinct human cytosolic sulfotransferase (ST), hST1B2, has been isolated from a human liver λZap cDNA library. The hST1B2 cDNA consists of 1144 bp and contains the coding region for a novel human cytosolic ST that has been termed hST1B2 on the basis of its sequence similarity to a rat sulfotransferase, ST1B1. The hST1B2 cDNA contains an 888-bp open reading frame that encodes a 296-amino acid protein with a calculated molecular mass of 34,897 Da. The hST1B2 cDNA also has a 127-bp 5′ untranslated region (UTR) and a 129-bp 3′-UTR, including a 22-bp poly(A)+ tract. The amino acid sequence of hST1B2 is 74%, 53%, 53%, 52%, 56%, and 34% identical to the amino acid sequences of rat ST1B1 and human P-PST-1, P-PST-2, M-PST, EST, and DHEA-ST, respectively. Enzymatically active hST1B2 was expressed in the bacterial expression vector pKK233–2 for kinetic characterization and in the bacterial expression vector pQE-31, which generates a histidine-tagged fusion protein for the generation of antibodies. Expressed hST1B2 sulfates small phenols such as 1-naphthol andp-nitrophenol and thyroid hormones, including 3,3′-diiodothyronine, triiodothyronine, reverse triiodothyronine, and thyroxine. No activity was detected when several steroids or dopamine were tested as substrates. High levels of hST1B2 message were detected by Northern blot analysis in RNA isolated from human liver, colon, small intestine, and blood leukocytes. Immunoblot analysis detected a protein with the same mass as expressed hST1B2 in several human tissues that also possessed hST1B2 message. These results indicate that a novel cytosolic ST is present in human tissues, which may have an important role in thyroid hormone and xenobiotic metabolism.
Footnotes
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Send reprint requests to: Charles N. Falany, Ph.D., Department of Pharmacology, 101 Volker Hall, 1670 University Boulevard, University of Alabama at Birmingham, Birmingham, AL 35294. E-mail:charles.falany{at}ccc.uab.edu
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This work was supported by National Institutes of Health Grant GM38953.
- Abbreviations:
- PAPS
- 3′-phosphoadenosine-5′-phosphosulfate
- ST
- sulfotransferase
- hDHEA-ST
- human dehydroepiandrosterone sulfotransferase
- PST
- phenol sulfotransferase
- hEST
- human estrogen sulfotransferase
- hP-PST-1
- human phenol-sulfating form of phenol sulfotransferase
- hM-PST
- human monoamine-sulfating form of phenol sulfotransferase
- E1
- estrone
- E2
- β-estradiol
- DHEA
- dehydroepiandrosterone
- T4
- 3,3′,5,5′-tetraiodothyronine (thyroxine)
- T3
- 3,5,3′-triiodothyronine
- rT3
- 3,3′,5′-triiodothyronine
- 3
- 5-T2, 3,5-diiodothyronine
- 3
- 3′-T2, 3,3′-diiodothyronine
- ID-I
- type I iodothyronine deiodinase
- PCR
- polymerase chain reaction
- pfu
- Pyrococcus furiosus
- bp
- base pair(s)
- UTR
- untranslated region
- SDS
- sodium dodecyl sulfate
- PAGE
- polyacrylamide gel electrophoresis
- TLC
- thin layer chromatography
- MBP
- maltose binding protein
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- Received April 16, 1997.
- Accepted October 24, 1997.
- The American Society for Pharmacology and Experimental Therapeutics
