Further characterization of human microsomal 3alpha-hydroxysteroid dehydrogenase.

Chetyrkin SV, Hu J, Gough WH, Dumaual N, Kedishvili NY
Arch Biochem Biophys. 2001 386 (1): 1-10

PMID: 11360992 · DOI:10.1006/abbi.2000.2203

This manuscript reports further characterization of the recently discovered human short-chain alcohol dehydrogenase, proposed to oxidize 3alpha-androstanediol to dihydrotestosterone in testis and prostate (M. G. Biswas and D. W. Russell, 1997, J. Biol. Chem. 272, 15959-15966). Enzyme expressed using the Baculovirus System localized in the microsomal fraction and catalyzed oxidation and reduction of the functional groups on steroids at carbons 3 and 17. Autoradiography assays revealed that the enzyme was most efficient as a 3alpha-hydroxysteroid oxidoreductase. High affinity of the enzyme for NADH (Km of 0.18 microM), lack of stereospecificity in the reductive direction, and poor efficiency for 3beta- versus 3alpha-hydroxyl oxidation could account for the observed transient accumulation of 3beta-stereoisomers in the oxidative reaction. Consistent with the 65% sequence identity with RoDH dehydrogenases, the enzyme oxidized all-trans-retinol with the Km value of 3.2 microM and Vmax value of 1.2 nmol/min per milligram microsomes. 13-cis-Retinol and all-trans-retinol bound to the cellular retinol-binding protein were not substrates. Neurosteroid allopregnanolone was a better substrate than all-trans-retinol with the Km and Vmax values of 0.24 microM and 14.7 nmol/min per milligram microsomes. Northern blot analysis revealed that the corresponding mRNA was present in adult human brain (caudate nucleus, amygdala, hippocampus, substantia nigra, thalamus) and spinal cord in addition to other tissues. The message was also detected in fetal lung, liver, and brain. Antibodies against the enzyme recognized a protein of approximately 35 kDa in the particulate fraction of human tissues. This study presents new information about enzymatic properties, substrate specificity, and tissue distribution of this enzyme, and provides a better insight into its possible physiological function(s).

MeSH Terms (26)

3-Hydroxysteroid Dehydrogenases Animals Baculoviridae Blotting, Northern Blotting, Western Brain Carbon Cell Line Cloning, Molecular Humans Insecta Kinetics Liver Lung Male Microsomes Models, Chemical Oxygen Prostate Recombinant Proteins RNA, Messenger Substrate Specificity Testis Time Factors Tissue Distribution Vitamin A

Connections (1)

This publication is referenced by other Labnodes entities:

Links