Cloning and expression of a novel sulfotransferase with unique substrate specificity from Bombyx mori.
We identified a cDNA encoding a putative cytosolic sulfotransferase (SULT) by searching the expressed sequence tag database of Bombyx mori, and subsequently obtained the full-length cDNA for this gene via rapid amplification of cDNA ends (RACE). We designated this gene bmST1, and showed by sequence analysis that it belongs to a novel SULT family. The tissue specificity of bmST1 mRNA expression was examined in fifth instar larvae by reverse transcriptase-polymerase chain reaction (RT-PCR), and transcripts were detectable in the silk gland, gut, fat body, and Malpighian tube. A recombinant form of bmST1 was then expressed using a gluthathione S-transferase (GST) gene fusion system, and it was purified from Escherichia coli. Purified bmST1 did not exhibit sulfating activity toward SULT substrates such as 4-nitrophenol, vanillin, hydroxysteroids, or monoamines. Surprisingly, however, recombinant bmST1 showed considerable activity toward 4-nitrocatechol and also gallate esters, although the catechins are not sulfated by this enzyme.
Hattori
Kenji
K
Kyoritsu University of Pharmacy, Tokyo, Japan.
Hirayama
Mio
M
Suzuki
Hiroko
H
Hamamoto
Hiroshi
H
Sekimizu
Kazuhisa
K
Tamura
Hiro-Omi
HO
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D012831:Bombyx / Q000201:enzymology*; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000096:biosynthesis / Q000235:genetics; D030541:Databases, Genetic; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013379:Substrate Specificity; D015238:Sulfotransferases / Q000096:biosynthesis* / Q000235:genetics* / Q000378:metabolism; D014018:Tissue Distribution
D018076:DNA, Complementary; D012333:RNA, Messenger; D015238:Sulfotransferases