CLONING, SEQUENCING AND EXPRESSION OF PSEUDOMONAS-TESTOSTERONI GENE ENCODING 3-ALPHA-HYDROXYSTEROID DEHYDROGENASE

We describe the cloning, sequencing and expression of the 3 alpha-hydr oxysteroid dehydrogenase (3 alpha-HSD) gene of Pseudomonas testosteron i. A genomic library of P. testosteroni total DNA constructed from Sau IIIA digests ligated to an lambda gtII vector was probed with a polycl onal antibody raised against purified enzyme. Subclones derived from a recombinant phage containing a 1746 bp insert were sequenced and foun d to contain an open reading frame of 696 bp that corresponds to a pro tein of 231 amino acid residues. A search for homologous proteins was performed. No similarity was observed when comparing 3 alpha-HSD with known members of the short-chain dehydrogenase family. However a small proteic fragment (80 amino acids) shows homology with the N-terminal sequence of bacterial L7/L12 ribosomal proteins.