Molecular cloning, expression, localisation and functional characterisation of a rabbit SULT1C2 sulfotransferase.
The importance of sulfotransferases in xenobiotic metabolism is gaining recognition. The gastrointestinal (GI) tract is a major portal of entry for many xenobiotics, yet little is known about the contribution of sulfotransferases to detoxication or bioactivation metabolism in these tissues. To this end, isolation and characterisation of sulfotransferases expressed in the stomach of rabbits was undertaken. A unique sulfotransferase cDNA (GenBank Accession No. AF026304) was isolated from a rabbit stomach cDNA library. This cDNA was 1439 base pairs (bp) long and has an open reading frame of 888 bp. On expression of the cDNA in both COS cells and E. coli, a protein molecular weight of 34 kDa was detected on SDS-PAGE. Immunoblotting using an antibody raised in goats against the bacterially expressed protein detected expression of the protein in GI tract tissues. The 34 kDa immunoreactive band was detected in rabbit GI tract tissues (stomach, duodenum, jejunum, ileum, colon, caecum and rectum), liver and kidneys, but not in the lungs (n = 3). The human ortholog (GenBank Accession No AF026303) of the rabbit enzyme was cloned from a human stomach cDNA library. These two enzymes share 84% amino acid sequence identity and have been termed 1C2 sulfotransferases. When functional and kinetic characterisation of the recombinant rabbit and human proteins was carried out using 16 known ST substrates, detectable sulfonation activity was observed only with p-nitrophenol (with Km values of 2.2 mM and 13.3 mM, respectively). In conclusion, we have identified a rabbit GI tract sulfotransferase belonging to a newly defined sulfotransferase subfamily.
Hehonah
N
N
Department of Physiology and Pharmacology, University of Queensland, Brisbane, Australia.
Zhu
X
X
Brix
L
L
Bolton-Grob
R
R
Barnett
A
A
Windmill
K
K
McManus
M
M
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D019556:COS Cells; D003001:Cloning, Molecular; D004064:Digestive System / Q000201:enzymology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004795:Enzyme Stability; D004926:Escherichia coli; D015971:Gene Expression Regulation, Enzymologic; D007700:Kinetics; D008969:Molecular Sequence Data; D009596:Nitrophenols / Q000494:pharmacology; D011817:Rabbits; D016415:Sequence Alignment; D015238:Sulfotransferases / Q000737:chemistry / Q000235:genetics*; D014162:Transfection
D004791:Enzyme Inhibitors; D009596:Nitrophenols; C015802:2,6-dichloro-4-nitrophenol; C513319:SULT1C2 protein, human; D015238:Sulfotransferases