Molecular cloning, expression, characterization, and ontogenic study of novel zebrafish cytosolic SULTs

By searching the expressed sequence tag database, five novel zebrafish SULT cDNAs were identified. The recombinant proteins of these SULTs, designated SULT1 ST7 and ST8, SULT2 ST4, and SULT3 ST1 and ST2, were expressed using the pGEX-2TK GST gene fusion system and purified from transformed E. coli cells. The enzymatic characterization of the purified proteins was performed. Purified SULT1 ST7 and ST8 exhibited strong sulfating-activities toward n-propyl gallate and polychlorinated biphenylols, among various endogenous and xenobiotic compounds tested as substrates. SULT2 ST4 displayed a high substrate specificity toward dehydroepiandrosterone (DHEA) and pregnenolone; whereas SULT3 ST1 and ST2 exhibited strong activities toward 17-estradiol and DHEA. The pH-dependence and kinetic constants of these enzymes were determined. Some of the known environmental estrogens and divalent metal cations exerted inhibitory effects on the sulfating activities of these SULTs. Developmental stage-dependent expression experiments revealed distinct patterns of expression of these SULTs during embryonic development and throughout the larval stage onto maturity. These results provide essential information pertaining to the use of the zebrafish as a model for systematic investigation on fundamental issues regarding the cytosolic SULTs.