Cholesterol oxidase (ChO) is an enzyme that usually gets involved in oxidation reactions in the cholesterol pathway. The cholesterol oxidase is an enzyme, most active on cholesterol molecules and converts cholesterol to cholesterol, that enzyme is the heart of cholesterol. This cholesterol metabolism is physiologically significant - bile acids and steroid hormones are cholesterol. In a normal environment, the role of cholesterol oxidase is less about degradation of cholestane skeleton substrates and more about cholesterol metabolism. In general, cleosteroid molecules (eg cholesterol) are skeleton substrates. These substrates participate in all kinds of bodily functions - from the architecture of cell membranes, to hormone production. Cholesterol oxidase does not direct these substrates toward degradation but engages in cholesterol's oxidation reaction in a number of metabolic chains.
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Cholesterol Oxidase
Cholesterol oxidase (ChO) is an enzyme that's mainly responsible for attacking cholesterol molecules, turning cholesterol into cholesterol. This enzyme is present in all microbes, plants, and animals, and it functions along the cholesterol metabolism. Cholesterol oxidase is an oxidase that allows cholesterol to be oxidized. It converts cholesterol into cholesterol and makes hydrogen peroxide and a ketone. This is the standard reaction bacteria and fungi use to use cholesterol as a carbon source. There are various uses of cholesterol oxidase in biotechnology and biochemistry. - for instance, to analyze and count cholesterol content, prepare cholesterol, etc. Furthermore, because cholesterol is selectively oxidized, cholesterol oxidase can also be used to create compounds like oxidized cholesterol.
Design of Cholesterol Oxidases
Cholesterol oxidases of various microbes have also been protein engineered and have been the focus of biotechnological speculation. The new putative cholesterol oxidase gene PsChO22 was recently identified from P simplex, one of the most popular steroid conversion bacteria, as it is highly resistant to organic solvents. PsChO is a new flavin-dependent enzyme that dehydrogenates the substrate of the cholestane backbone at C3. Further, structural comparisons of WT and mutants using MD revealed substrate preference of PsChO. We obtained more catalytically active enzymes by using molecular modelling with mutagenesis and biological experiments to generate PsChO mutants that were more specific and active on substrates.
Fig. 1 Structural analysis of the PsChO-substrate complex (Qin, H., et al. 2017).
Characteristics of Active Site
The site for the non-covalent form of ChO family enzymes7 lies below the isooxazine of FAD. Three residues, H431, E346, and N469 make up the catalytic triad. Residue H431 is a universal base catalyst to pulls a proton from the C3-OH of the steroid substrate during oxidation and a universal acid to fixate the dienolate intermediate of the isomerism 23. E346 is found near the steroid's C4 -proton. E346 is the starting material of the proton transfer and the isomerization step 13. N469 regulates flavin oxidation and also its redox potential via the link between the flavin side chain and the ring of FAD 26. The water molecule at the active site is joined by a hydrogen network between the catalytic residues H431, E346, and N469 and the substrate's C3-OH.
Cholestane Substrates
The cholesterol is a fundamental lipid (synthetic) substance, with many biological functions. Cholesterol is dehydrated and used by the body as a byproduct of enzymes like cholesterol dehydrogenase, cholesterol esterase, and cholesterol esterase. Ester chemicals are cholesterol, which is made of cholesterol and fatty acids. As a food, cholesterol esterase in the gut hydrolyses it into free cholesterol and free fatty acids for absorption. Cholesterol oxidase reacts to oxidize cholesterol and transforms cholesterol into cholestenol (heptane peroxide, ketone). Cells have a complicated cholesterol metabolism, and cholesterol dissolves thanks to the work of all these enzymes because it enters the cellular cycle. These pathways in the metabolic cycle need to be in balance and controlled for normal functioning of the body, as well as for the stability of cholesterol levels.
Promoted Cholestane Substrate Decomposition
Cholesterol oxidase oxidises and isomerizes cholestane substrates, whereupon a hydroxyl group is introduced on the C3 side. The Pimelobacter simplex cholesterol oxidase (PsChO) was rationally designed. Mutations of V64 and F70 made the catalytic activity towards cholestane substrates better. Through a combination of molecular dynamics simulations and structure-activity correlations, we discovered that V64C and F70V raised the free energy at which the PsChO mutants bound to cholesterol. F70V and V64C can also change the lollipops L56-P77, K45-P49, and L350-E354 around in the active site. They open up the substrate binding cavity and reduce the steric interference with the substrate to favor C17 hydrophobic substrates over long side chain substrates.
Conclusion
Modification of cholesterol oxidase: Some modification or modification of the enzyme to alter its activity, stability, or other biological attributes and hence modify its biological activity. This can be done by any number of means like phosphorylation, methylation, glycosylation, and methylation. Changes to cholesterol oxidase can modulate how active or stable cholesterol oxidase is in certain conditions and also oxidize more substrates or substrates of different structures.
References
- Qin, H., et al. Rational design of cholesterol oxidase for efficient bioresolution of cholestane skeleton substrates. Scientific reports. 2017, 7(1): 16375.
- Aggarwal, V., et al. Amperometric determination of serum total cholesterol with nanoparticles of cholesterol esterase and cholesterol oxidase. Analytical Biochemistry. 2016, 500: 6-11.
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