7β-hydroxysteroid dehydrogenases and their use
AssigneePharmaZell GmbH
InventorsLou Liu, Rolf Schmid, Arno Aigner
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Abstract
The invention relates to novel 7β-hydroxysteroid dehydrogenases which are obtainable from bacteria of the genus Collinsella, especially of the strain Collinsella aerofaciens, to the sequences encoding said enzymes, to methods for producing said enzymes and to their use in the enzymatic conversion of cholic acid compounds, and especially in the production of ursodeoxycholic acid (UDCS). The invention also relates to novel methods for the synthesis UDCS.
Claims22
An isolated recombinant 7β-hydroxysteroid dehydrogenase (7β-HSDH) comprising an amino acid sequence according to SEQ ID NO:
- 2 or an amino acid sequence having only conservative amino acid substitutions as compared to SEQ ID NO:
- 2 and having at least 95% sequence identity to SEQ ID NO:
- 2 and a terminal His-Tag sequence, wherein the recombinant 7β-HSDH has a molecular weight, determined by gel filtration, in the range of 53 to 60 kDa, and wherein the recombinant 7β-HSDH is at least capable of catalyzing stereospecific reduction of a 7-ketosteroid to the corresponding 7β-hydroxysteroid.
An isolated recombinant 7β-hydroxysteroid dehydrogenase (7β-HSDH) comprising an amino acid sequence according to SEQ ID NO:
- 2 or an amino acid sequence having only conservative amino acid substitutions as compared to SEQ ID NO:
- 2 and having at least 95% sequence identity to SEQ ID NO:
- 2 and a terminal His-Tag sequence, wherein the recombinant 7β-HSDH has a molecular weight, determined by gel filtration, in the range of 53 to 60 kDa and is at least capable of catalyzing stereospecific reduction of a 7-ketosteroid to the corresponding 7β-hydroxysteroid, and wherein the recombinant 7β-HSDH is prepared by a process comprising the steps of:
- cloning a gene comprising the nucleic acid sequence of SEQ ID NO:1 or a nucleic acid sequence having at least 95% sequence homology with SEQ ID NO:1 and a nucleic acid sequence encoding a His-Tag;
- expressing the cloned gene in a bacterial host;
- and
- isolating the expressed protein.
Dependencies are inferred from the claim text.