Combination of C(17) sphingoid base homologues and mass spectrometry analysis as a new approach to study sphingolipid metabolism

Methods Enzymol. 2007;434:233-41. doi: 10.1016/S0076-6879(07)34012-3.

Abstract

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.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Animals
  • Chromatography, High Pressure Liquid / methods
  • Fatty Acids / isolation & purification
  • Fatty Acids / metabolism
  • Kinetics
  • Mass Spectrometry / methods
  • Oxidoreductases / isolation & purification
  • Oxidoreductases / metabolism*
  • Phosphotransferases (Alcohol Group Acceptor) / isolation & purification
  • Phosphotransferases (Alcohol Group Acceptor) / metabolism*
  • Sphingolipids / isolation & purification*
  • Sphingolipids / metabolism*

Substances

  • Fatty Acids
  • Sphingolipids
  • Oxidoreductases
  • dihydroceramide desaturase
  • Phosphotransferases (Alcohol Group Acceptor)
  • sphingosine kinase