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  • A novel alkyne cholesterol to trace cellular cholesterol metabolism and localization.

A novel alkyne cholesterol to trace cellular cholesterol metabolism and localization.

Journal of lipid research (2013-12-18)
Kristina Hofmann, Christoph Thiele, Hans-Frieder Schött, Anne Gaebler, Mario Schoene, Yuriy Kiver, Silvia Friedrichs, Dieter Lütjohann, Lars Kuerschner
ABSTRACT

Cholesterol is an important lipid of mammalian cells and plays a fundamental role in many biological processes. Its concentration in the various cellular membranes differs and is tightly regulated. Here, we present a novel alkyne cholesterol analog suitable for tracing both cholesterol metabolism and localization. This probe can be detected by click chemistry employing various reporter azides. Alkyne cholesterol is accepted by cellular enzymes from different biological species (Brevibacterium, yeast, rat, human) and these enzymes include cholesterol oxidases, hydroxylases, and acyl transferases that generate the expected metabolites in in vitro and in vivo assays. Using fluorescence microscopy, we studied the distribution of cholesterol at subcellular resolution, detecting the lipid in the Golgi and at the plasma membrane, but also in the endoplasmic reticulum and mitochondria. In summary, alkyne cholesterol represents a versatile, sensitive, and easy-to-use tool for tracking cellular cholesterol metabolism and localization as it allows for manifold detection methods including mass spectrometry, thin-layer chromatography/fluorography, and fluorescence microscopy.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
N-(3-Azidopropyl)biotinamide, ≥95%
Avanti
alkyne-cholesterol, 27-norcholest-5-en-25-yn-3β-ol, powder