Open AccessCholesterol oxidase from Rhodococcus erythropolis with high specificity toward β-cholestanol and pytosterols
Author(s) -
Noriyuki Doukyu,
Makoto Ishikawa
Publication year - 2020
Publication title -
plos one
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.99
H-Index - 332
ISSN - 1932-6203
DOI - 10.1371/journal.pone.0241126
Subject(s) - cholesterol oxidase , cholestanol , biochemistry , sterol , biology , escherichia coli , cholesterol , enzyme , rhodococcus , gene , chemistry
Two genes ( choRI and choRII ) encoding cholesterol oxidases belonging to the vanillyl-alcohol oxidase (VAO) family were cloned on the basis of putative cholesterol oxidase gene sequences in the genome sequence data of Rhodococcus erythropolis PR4. The genes corresponding to the mature enzymes were cloned in a pET vector and expressed in Escherichia coli . The two cholesterol oxidases produced from the recombinant E . coli were purified to examine their properties. The amino acid sequence of ChoRI showed significant similarity (57%) to that of ChoRII. ChoRII was more stable than ChoRI in terms of pH and thermal stability. The substrate specificities of these enzymes differed distinctively from one another. Interestingly, the activities of ChoRII toward β-cholestanol, β-sitosterol, and stigmasterol were 2.4-, 2.1-, and 1.7-fold higher, respectively, than those of cholesterol. No cholesterol oxidases with high activity toward these sterols have been reported so far. The cholesterol oxidation products from these two enzymes also differed. ChoRI and ChoRII oxidized cholesterol to form cholest-4-en-3-one and 6β-hydroperoxycholest-4-en-3-one, respectively.