Combination of C(17) sphingoid base homologues and mass spectrometry analysis as a new approach to study sphingolipid metabolism.
In recent years, sphingolipid metabolites ceramide, sphingosine, and sphingosine-1-phosphate have emerged as important second messengers in addition to their role as precursors of biomembrane components. The investigation of these sphingolipid metabolites requires the development of new, more sensitive methods for assaying the enzymes involved in their production. This chapter describes the utilization of mass spectrometry technology in combination with nonnaturally occurring C(17) sphingoid bases in the in vitro assays of two of the enzymes of the sphingolipid pathway, ceramide synthase and sphingosine kinase. These new in vitro methods provide high sensitivity and extreme accuracy even when crude extracts are used as enzyme sources.
Spassieva
Stefka
S
Department of Medicine, Medical University of South Carolina, Charleston, South Carolina, USA.
Bielawski
Jacek
J
Anelli
Viviana
V
Obeid
Lina M
LM
D000818:Animals; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D005227:Fatty Acids / Q000302:isolation & purification / Q000378:metabolism; D007700:Kinetics; D013058:Mass Spectrometry / Q000379:methods; D010088:Oxidoreductases / Q000302:isolation & purification / Q000378:metabolism*; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000302:isolation & purification / Q000378:metabolism*; D013107:Sphingolipids / Q000302:isolation & purification* / Q000378:metabolism*; D000099157:Sphingosine Kinase
D005227:Fatty Acids; D010088:Oxidoreductases; D017853:Phosphotransferases (Alcohol Group Acceptor); D013107:Sphingolipids; D000099157:Sphingosine Kinase; C108138:dihydroceramide desaturase