MOLECULAR-CLONING OF TESTICULAR 20-ALPHA-HYDROXYSTEROID DEHYDROGENASE - IDENTITY WITH ALDOSE REDUCTASE

Complementary DNA (cDNA) clones encoding bovine testicular 20alpha-hydroxysteroid dehydrogenase (20alpha-HSD) have been isolated from a bovine testicular lambdagt11 library using polyclonal antibodies against 20alpha-HSD and DNA probe hybridization. Nucleotide sequencing of three independently isolated clones was used to establish a composite cDNA sequence that encodes the enzyme. It contains a coding sequence of 921 nucleotides, a stop codon, and a 264-nucleotide 3'-noncoding segment which allowed deduction of the amino acid sequence of the enzyme. A computer homology search of the 20alpha-HSD cDNA performed against the GenBank DNA sequence database revealed it to be identical with bovine lens aldose reductase (alditol:NADPH oxidoreductase; EC 1.1.1.21), and a literature search reveals the deduced amino acid sequence to be identical with that reported for the bovine enzyme. Sequences obtained from the N-terminus of purified testicular 20alpha-HSD and from random peptides obtained by treatment with endopeptidase Lys-C are all identical with regions of the deduced amino acid sequence of 20alpha-HSD and/or the published sequence of aldose reductase. Further, the enzyme purified to homogeneity by following activity with 17-hydroxyprogesterone as a substrate was shown to reduce glucose, glyceraldehyde, and benzaldehyde (all classic aldose reductase substrates). Finally, 17-hydroxyprogesterone inhibited the reduction of benzaldehyde and glyceraldehyde. Because aldose reductase has been implicated in the etiology of diabetic complications, acceptance of steroid substrates may offer new implications for therapy.