Applied Microbiology and Biotechnology

, Volume 85, Issue 6, pp 1831–1838

Cholesterol oxidase ChoL is a critical enzyme that catalyzes the conversion of diosgenin to 4-ene-3-keto steroids in Streptomyces virginiae IBL-14

Biotechnologically Relevant Enzymes and Proteins

Abstract

Diosgenin transformation was studied in Streptomyces virginiae IBL-14, a soil-dwelling bacterium with diosgenin-degrading capacity. All of the derivatives isolated were identified as 4-ene-3-keto steroids. We cloned ChoL, a fragment of a cholesterol oxidase from S. virginiae IBL-14, and used gene-disruption techniques to determine its function in the oxidation of diosgenin to 4-ene-3-keto steroids. Subsequently, the entire open reading frame of ChoL was cloned by chromosome walking, and the His6-tagged recombinant protein was overproduced, purified, and characterized. ChoL consisted of 1,629 nucleotides that encoded a protein of 542 amino acids, including a 34-residue putative signal peptide at the N-terminal. ChoL showed 85% amino acid similarity to ChoA from Streptomyces sp. SA-COO. This enzyme can also oxidize other steroids such as cholesterol, sitosterol, and dehydroepiandrosterone, which showed higher affinity (Km = 0.195 mM) to diosgenin. The catalytic properties of this enzyme indicate that it may be useful in diosgenin transformation, degradation, and assay.

Keywords

Cholesterol oxidase Streptomyces virginiae IBL-14 Diosgenin Diosgenone 4-ene-3-keto steroids 

Supplementary material

253_2009_2188_MOESM1_ESM.doc (424 kb)
ESM 1(DOC 424 kb)

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Copyright information

© Springer-Verlag 2009

Authors and Affiliations

  1. 1.State Key Laboratory of Bioreactor Engineering, Newworld Institute of BiotechnologyEast China University of Science and TechnologyShanghaiChina
  2. 2.East China University of Science and TechnologyShanghaiChina

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