Estradiol 17 beta-dehydrogenase: full enzymatic activity in the absence of zinc.
The precise catalytic mechanism of the steroid interconverting enzyme, human placental estradiol 17 beta-dehydrogenase (EC 1.1.1.62, estradiol-17 beta:NAD+ 17-oxidoreductase), is not known. Two general models for the catalytic mechanism of dehydrogenases have been defined. One model requires Zn2+ metal for the catalytic event, as has been shown for horse liver alcohol dehydrogenase (EC 1.1.1.1, alcohol:NAD+ oxidoreductase). Another model has been demonstrated for the 2-hydroxy acid dehydrogenases in which histidine residues are necessary for enzyme activity, without participation of a metal ion. In order to define which mechanism might be operative for the placental enzyme, it became important to determine whether Zn2+, or another metal ion, is associated with the macromolecule. Several homogeneous enzyme preparations, having protein concentrations from 5-80 microM, were extensively dialyzed in a buffer containing EDTA. Atomic absorption analysis of each sample demonstrated that no Zn2+ was present, although the enzymatic activity was maintained. In addition, there was no significant detection of Mg2+ or Mn2+ above background levels. When the isolated enzyme was dialyzed against buffer containing added 0.01-20 microM ZnCl2, no increase in specific activity of the enzyme was seen. The data indicate that the presence of zinc is not required for the catalytic event. These results, together with our previous affinity-labeling studies, which demonstrate a histidine residue in the catalytic region of the active site, allow us to propose that the catalytic mechanism of the human placental estradiol 17 beta-dehydrogenase is similar to that of the 2-hydroxy acid dehydrogenases.
Murdock
G L
GL
Department of Obstetrics/Gynecology, Washington University School of Medicine, St. Louis, MO.
Pineda
J
J
Nagorsky
N
N
Lawrence
S S
SS
Heritage
R
R
Warren
J C
JC
D000595:Amino Acid Sequence; D001665:Binding Sites; D002846:Chromatography, Affinity; D004960:Estradiol Dehydrogenases / Q000302:isolation & purification / Q000378:metabolism*; D005260:Female; D006801:Humans; D007700:Kinetics; D008958:Models, Molecular; D010920:Placenta / Q000201:enzymology; D011247:Pregnancy; D011487:Protein Conformation; D013054:Spectrophotometry, Atomic; D015032:Zinc / Q000032:analysis / Q000494:pharmacology*
D004960:Estradiol Dehydrogenases; D015032:Zinc