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Preparation and Properties of a Cholesterol Oxidase from Nocardia sp. and Its Application to the Enzymatic Assay of Total Cholesterol in Serum

Clinical Chemistry · 1973 · Vol. 19(12) · pp. 1350–1356

W. Richmond✉(Medical Research Council)

Abstract

Abstract I describe the characterization, extraction, and purification of a cholesterol:oxygen oxidoreductase (EC 1.1.3.6) from Nocardia sp. This enzyme catalyzes oxidation of cholesterol to Δ4-choIestenone, with production of hydrogen peroxide. It is very stable, active over a wide pH range, and has a Km of 1.4 x 10-5 mol/ liter. It is highly specific for Δ4- or Δ5-3β-hydroxycholestanes, and may be applied to the assay of serum total cholesterol. In the procedure presented here, hydrogen peroxide is measured by reaction with quadrivalent titanium and xylenol orange. This constitutes a one-enzyme assay with stable reagents, which does not require protein precipitation and is not subject to interference from hemoglobin or bilirubin.

Mass Spectrometry Techniques and Applications Hemoglobin structure and function Receptor Mechanisms and SignalingCholesterol oxidaseXylenol orangeHydrogen peroxideChemistryEnzymeChromatographyHemoglobinCholesterolOxidoreductaseReagent

MeSH terms

AcetoneAlcohol OxidoreductasesAlcoholsAndrostenolsBilirubinCatalaseCell FractionationCholanesCholestenesCholesterolEvaluation Studies as TopicHemoglobinsHumansKineticsMethods

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