Sphingosine 1-phosphate lyase enzyme assay using a BODIPY-labeled substrate
- Cancer Biology Lab, Children's Hospital Oakland Research Institute, 5700 Martin Luther King Jr. Way, Oakland, CA 94609 (United States)
- Department of Chemistry and Biochemistry, Queens College, City University of New York, Flushing, NY 11367-1597 (United States)
Sphingosine 1-phosphate lyase (SPL) is responsible for the irreversible catabolism of sphingosine 1-phosphate, which signals through five membrane receptors to mediate cell stress responses, angiogenesis, and lymphocyte trafficking. The standard assay for SPL activity utilizes a radioactive dihydrosphingosine 1-phosphate substrate and is expensive and cumbersome. In this study, we describe an SPL assay that employs an {omega}-labeled BODIPY-sphingosine 1-phosphate substrate, allowing fluorescent product detection by HPLC and incorporating advantages of the BODIPY fluorophore. The major aldehyde product is confirmed by reaction with 2,4-dinitrophenylhydrazine. The SPL-catalyzed reaction is linear over a 30 min time period and yields a K{sub m} of 35 {mu}M for BODIPY-sphingosine 1-phosphate.
- OSTI ID:
- 21255919
- Journal Information:
- Biochemical and Biophysical Research Communications, Vol. 380, Issue 2; Other Information: DOI: 10.1016/j.bbrc.2009.01.106; PII: S0006-291X(09)00128-4; Copyright (c) 2009 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved; Country of input: International Atomic Energy Agency (IAEA); ISSN 0006-291X
- Country of Publication:
- United States
- Language:
- English
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