Journal
FEBS LETTERS
Volume 581, Issue 27, Pages 5289-5294Publisher
ELSEVIER SCIENCE BV
DOI: 10.1016/j.febslet.2007.10.018
Keywords
lipid; sphingolipid; ceramide; acyl CoA; sphinganine; assay
Funding
- NCI NIH HHS [R01CA085704, R01CA052462, R01CA105125] Funding Source: Medline
- NIGMS NIH HHS [GM076217] Funding Source: Medline
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Ceramide is a key metabolite in the pathway of sphingolipid biosynthesis. In mammals, ceramide is synthesized by N-acylation of a sphingoid long-chain base by a family of ceramide synthases (CerS), each of which displays a high specificity towards acyl CoAs of different chain lengths. We now optimize a previously-described assay for measuring CerS activity for use upon over-expression of mammalian CerS, and using these conditions, establish the Km value of each CerS towards sphinganine. Remarkably, the Km values towards sphinganine are all similar, ranging from 2 to 5 mu M, even for CerS proteins that are able to use more than one acyl CoA for ceramide synthesis (i.e. CerS4). The availability of this assay will permit further accurate characterization of the kinetic parameters of mammalian CerS proteins. (C) 2007 Federation of European Biochemical Societies. Published by Elsevier B. V. All rights reserved.
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