cDNA cloning, expression, and functional characterization of a zebrafish SULT1 cytosolic sulfotransferase.
Using the reverse transcriptase-polymerase chain reaction technique, a full-length cDNA encoding a novel zebrafish sulfotransferase was cloned and sequenced. Sequence analysis indicated that this zebrafish sulfotransferase belongs to the SULT1 cytosolic sulfotransferase gene family. The recombinant form of the zebrafish sulfotransferase, purified from Escherichia coli cells, displayed sulfating activities toward a number of endogenous compounds, in particular dopamine and thyroid hormones, in addition to xenobiotics including some flavonoids, isoflavonoids, and other phenolic compounds. The zebrafish sulfotransferase exhibited substrate dependence in pH optimum. In comparison with those determined with dopamine as substrate, the zebrafish sulfotransferase displayed much lower K(m) and higher V(max) with n-propyl gallate as substrate. A thermostability experiment revealed the enzyme to be relatively stable over a temperature range between 20 and 43 degrees C. Among 10 divalent metal cations tested, Hg(2+), Co(2+), Zn(2+), Cd(2+), Cu(2+), and Pb(2+) exhibited dramatic inhibitory effects on the activity of the zebrafish sulfotransferase.
Sugahara
Takuya
T
Biomedical Research Center, The University of Texas Health Center, Tyler 75708, USA.
Liu
Chau-Ching
CC
Carter
Glendora
G
Govind Pai
T
T
Liu
Ming-Cheh
MC
D000818:Animals; D001483:Base Sequence; D002413:Cations, Divalent / Q000737:chemistry; D003001:Cloning, Molecular / Q000379:methods; D003600:Cytosol / Q000201:enzymology / Q000378:metabolism; D018076:DNA, Complementary / Q000235:genetics*; D004789:Enzyme Activation; D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics / Q000302:isolation & purification; D015971:Gene Expression Regulation, Enzymologic; D006863:Hydrogen-Ion Concentration; D008670:Metals / Q000737:chemistry; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D015238:Sulfotransferases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D013696:Temperature; D015027:Zebrafish / Q000235:genetics* / Q000378:metabolism*
D002413:Cations, Divalent; D018076:DNA, Complementary; D008670:Metals; D011993:Recombinant Fusion Proteins; D015238:Sulfotransferases