Catalytic differences between porcine blastocyst and placental aromatase isozymes.
Two isozymes of porcine aromatase, the placental and the blastocyst forms, were expressed in CHO cells using the mammalian cell transfection method. Using an 'in-cell' assay (a 3H-water release method), catalytic parameters of the porcine placental aromatase were found to be very similar to those of the human enzyme; however, the activity of the blastocyst isozyme was found to be one-thirtieth that of the placental isozyme. Product isolation assay (using testosterone as the substrate) revealed that the major steroid products were 17beta-estradiol and 19-nortestosterone. The product ratio of estradiol/19-nortestosterone was found to be 94 : 6 for the porcine placental form, 6 : 94 for the porcine blastocyst form, and 92 : 8 for the human wild-type aromatase. Therefore, the porcine blastocyst aromatase isozyme catalyzes mainly androgen 19-desmethylation rather than aromatization. In addition, inhibition profile analyses on the placental and blastocyst isozymes were performed using three steroidal inhibitors [4-hydroxyandro-stenedione (4-OHA), 7alpha-(4'-amino)phenylthio-1, 4-androstandiene-3,17-dione (7alpha-APTADD), and bridge (2, 19-methyleneoxy) androstene-3,17-dione (MDL 101,003)], and four nonsteroidal inhibitors [aminoglutethimide (AG), CGS 20267, ICI D1033, and vorozole (R83842)]. While the two isozymes of porcine aromatase share 93% amino-acid sequence identity, our results indicate that the two porcine aromatase isozymes have distinct responses to various aromatase inhibitors.
Kao
Y C
YC
Division of Immunology, Beckman Research Institute of the City of Hope, Duarte, CA, USA.
Higashiyama
T
T
Sun
X
X
Okubo
T
T
Yarborough
C
C
Choi
I
I
Osawa
Y
Y
Simmen
F A
FA
Chen
S
S
D000818:Animals; D001141:Aromatase / Q000378:metabolism*; D001755:Blastocyst / Q000201:enzymology*; D016466:CHO Cells; D002384:Catalysis; D006224:Cricetinae; D005260:Female; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism; D007700:Kinetics; D010920:Placenta / Q000201:enzymology*; D011247:Pregnancy; D011994:Recombinant Proteins / Q000378:metabolism; D013552:Swine; D014162:Transfection
D007527:Isoenzymes; D011994:Recombinant Proteins; D001141:Aromatase