Stable carbon isotope discrimination by human 3-hydroxy-3-methylglutaryl-coenzyme A reductase

• Main Authors: Svena M. Lüdke, Ulrich Flenker, Wilhelm Schänzer, Dietmar Schomburg
• Format: Article
• Language: English
• Published: Elsevier 2008-12-01
• Series: Journal of Lipid Research
• Subjects: mevalonate pathway; steroids; kinetic isotope effect; isotope ratio mass spectrometry; curve fitting; doping control
• Online Access: http://www.sciencedirect.com/science/article/pii/S0022227520345922
• ISSN: 0022-2275

The aim of this study was to investigate the possible existence and magnitude of stable carbon isotope discrimination by human 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGR). The catalytic portion of HMGR was expressed and purified. The reaction product mevalonate was lactonized and extracted from the reaction mixture by a solid-phase extraction protocol. Stable carbon isotope ratios of mevalonolactone (MVL) were analyzed by gas chromatography-combustion-isotope ratio mass spectrometry. An average fractionation factor 12k/13k of 1.0031 ± 0.0004 for all carbon atoms contained in MVL was estimated by the method of internal competition. The value was calculated by nonlinear curve fitting, where the ratio 13C/12C of MVL was plotted versus the fraction of reaction.