Cloning, DNA sequencing and expression of (3-17)beta hydroxysteroid dehydrogenase from Pseudomonas testosteroni.
We describe the cloning, sequencing and overexpression of the (3-17)beta hydroxysteroid dehydrogenase gene of Pseudomonas testosteroni. A genomic library of Ps. testosteroni total DNA constructed from SauIIIA digests ligated to a lambda gt11 vector was probed with polyclonal antibody raised against purified enzyme. Subclones derived from a recombinant phage containing a 2661-base-pair insert were sequenced and found to contain an open reading frame of 765 base pairs that corresponds to a protein of 254 amino acid residues. A 1492-base-pair fragment was inserted into pBR322 plasmid vector and used to construct a strain of E. coli HB101 that overexpressed the steroid dehydrogenase gene.
Abalain
J H
JH
Laboratoire de Biochimie, Faculté de Médecine, Brest, France.
Di Stefano
S
S
Amet
Y
Y
Quemener
E
E
Abalain-Colloc
M L
ML
Floch
H H
HH
D015067:17-Hydroxysteroid Dehydrogenases / Q000235:genetics* / Q000378:metabolism; D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004269:DNA, Bacterial; D004926:Escherichia coli; D015964:Gene Expression Regulation, Bacterial; D015971:Gene Expression Regulation, Enzymologic; D015151:Immunoblotting; D008969:Molecular Sequence Data; D011549:Pseudomonas / Q000201:enzymology*; D015183:Restriction Mapping
D004269:DNA, Bacterial; D015067:17-Hydroxysteroid Dehydrogenases; C044256:3 (or 17)-beta-hydroxysteroid dehydrogenase