Abstract
Cholesterol oxidase is an alcohol oxidoreductase flavoprotein with wide biotechnological applications. The current work describes the isolation of a potential cholesterol oxidase producing streptomycete from Egyptian soil. The isolated strain produced cholesterol oxidase in submerged culture using a medium containing glucose, yeast extract, malt extract, and CaCO3 with the addition of cholesterol as an inducer. The isolated strain was identified as Streptomyces rochei NAM-19 based on 16S rRNA sequencing and phylogeny. Optimization of cholesterol oxidase production has been carried out using response surface methodology. The Plackett-Burman design method was used to evaluate the significant components of the production medium followed by Box-Behnken experimental design to locate the true optimal concentrations, which are significantly affecting enzyme production. Results showed that the predicted enzyme response could be closely correlated with the experimentally obtained production. Furthermore, the applied optimization strategy increased volumetric enzyme production by 2.55 times (65.1 U/mL) the initial production obtained before medium optimization (25.5 U/mL).
Highlights
Cholesterol oxidase, CO (EC. 1.1.3.6) enzyme is a monomeric oxidoreductase flavoenzyme, which catalyzes the oxidation of cholesterol to cholesterone and hydrogen peroxide
CO enzyme-producing microbial strains were isolated from different soil samples collected from El-Giza Governorate, Giza, Egypt. e isolation medium is composed mainly of mineral agar screening medium supplemented with cholesterol as the sole carbon source for growth
BLAST software was used to compare the partial nucleotide sequence of the 16S rRNA gene of the CO-producing isolate with nucleotide databases found in NCBI webserver
Summary
Cholesterol oxidase, CO (EC. 1.1.3.6) enzyme is a monomeric oxidoreductase flavoenzyme, which catalyzes the oxidation of cholesterol to cholesterone and hydrogen peroxide. CO enzyme biosensors have found applications in the detection of cholesterol level in various samples [5] and have scientific importance in investigating cell membrane interactions with cholesterol [6]. CO enzyme has been produced by different microorganisms in submerged cultures, e.g., Arthrobacter sp., Pseudomonas sp., Rhodococcus sp., Mycobacterium sp., Streptomyces sp., Nocardia, and Streptoverticillium [7, 8]. Since CO enzyme has many applications in food and medical sectors, it is of great importance that the producing organisms should have the GRAS (generally regarded as safe) status [10]. Search for new and safe microorganisms capable of producing such an industrially important enzyme has continued.
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