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Báo cáo khoa học: Sulfation of hydroxychlorobiphenyls Molecular cloning, expression, and functional characterization of zebrafish SULT1 sulfotransferases
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As a first step toward developing a zebrafish model for investigating the role of sulfation in counteracting environ-mental estrogenic chemicals, we have embarked on the identification and characterization of cytosolic sulfotrans-ferases (STs) in zebrafish. By searching the zebrafish expressed sequence tag database, we have identified two cDNA clones encoding putative cytosolic STs. These two zebrafish ST cDNAs were isolated and subjected to nuc-leotide sequencing. Sequence data revealed that the two zebrafishSTs are highlyhomologous, being 82%identical in their amino acid sequences | Eur. J. Biochem. 270 2404-2411 2003 FEBS 2003 doi 10.1046 j 1432-1033.2003.03608.x Sulfation of hydroxychlorobiphenyls Molecular cloning expression and functional characterization of zebrafish SULT1 sulfotransferases Takuva Suaahara1. Chau-China Liu2 T. Govind Pai1. Paul Collodi3 Masahito Suiko1. Yoichi Sakakibara1 . Kazuo Nishiyama1 and Ming-Cheh Liu1 1 Biomedical Research Center The University of Texas Health Center Tyler Texas USA department of Medicine University of Pittsburgh School of Medicine Pittsburgh Pennsylvania 3Department of Animal Sciences Purdue University West Lafayette Illinois USA As a first step toward developing a zebrafish model for investigating the role of sulfation in counteracting environmental estrogenic chemicals we have embarked on the identification and characterization of cytosolic sulfotransferases STs in zebrafish. By searching the zebrafish expressed sequence tag database we have identified two cDNA clones encoding putative cytosolic STs. These two zebrafish ST cDNAs were isolated and subjected to nucleotide sequencing. Sequence data revealed that the two zebrafish STs are highly homologous being w 82 identical in their amino acid sequences. Both of them display w 50 amino acid sequence identity to human SULT1A1 rat SULT1A1 and mouse SULT1C1 ST. These two zebrafish STs therefore appear to belong to the SULT1 cytosolic ST gene family. Recombinant zebrafish STs designated SULT1 STs 1 and 2 expressed using the pGEX-2TK prokaryotic expression system and purified from transformed Escheri chia coli cells migrated as w 35 kDa proteins on SDS PAGE. Purified zebrafish SULT1 STs 1 and 2 displayed differential sulfating activities toward a number of endogenous compounds and xenobiotics including hydroxychlorobiphenyls. Kinetic constants of the two enzymes toward two representative hydroxychlorobiphenyls 3-chloro-4-biphenylol and 3 3 5 5 -tetrachloro-4 4 -biphenyldiol and 3 3 5-triiodo-L-thyronine were determined. A thermostability experiment .