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Leukotriene formation by purified 5-lipoxygenase from rat basophilic leukemia cells

Conference · · Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States)
OSTI ID:5096401
; ; ; ;

Arachidonate 5-lipoxygenase (5-LO) from rat basophilic leukemia (RBL-1) cell high speed (105,000 x g for 60 min) supernatants was purified to electrophoretic homogeneity by gel filtration and anion-exchange protein-high pressure liquid chromatography (HPLC). The 5-LO rapidly converted (/sup 14/C)arachidonate at 20/sup 0/C to (/sup 14/C)5-hydroperoxyeicosate-traenoic acid (HPETE) as determined by reversed phase-HPLC, scanning spectrophotometry and radiochemical detection. In addition, 5-LO converted both 5-HPETE and arachidonate to 5,12-dihydroxyeicosatetraenoic acids (diHETEs). The 5,12-diHETEs were identified as 6-trans-leukotriene (LT) B4 and 6-trans-12-epi-LTB4 as determined by reversed phase HPLC, scanning spectrophotometry and gas chromatography-mass spectrometry. These data indicate that the RBL-1 5-LO and LTA4 synthetase activities reside on the same protein and that it catalyzes the bioconversion of arachidonate to not only 5-HPETE but also to LTA4. The results suggest that a critical regulatory step in LT biosynthesis in mammalian systems involves the intricate coupling of the enzymes 5-LO and LTA4 synthetase and the interactions of their respective cofactors, substrates and reaction products.

Research Organization:
Smith Kline and French Labs., Philadelphia, PA
OSTI ID:
5096401
Report Number(s):
CONF-8606151-
Journal Information:
Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States), Journal Name: Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States) Vol. 45:6; ISSN FEPRA
Country of Publication:
United States
Language:
English

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Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
ANIMAL CELLS
ANIMALS
ARACHIDONIC ACID
BIOSYNTHESIS
CARBON 14 COMPOUNDS
CARBOXYLIC ACIDS
CHROMATOGRAPHY
DISEASES
ELECTROPHORESIS
ENZYMES
GAS CHROMATOGRAPHY
HEMIC DISEASES
LABELLED COMPOUNDS
LEUKEMIA
LIQUID COLUMN CHROMATOGRAPHY
MAMMALS
MASS SPECTROSCOPY
MONOCARBOXYLIC ACIDS
NEOPLASMS
ORGANIC ACIDS
ORGANIC COMPOUNDS
OXIDOREDUCTASES
OXYGENASES
PURIFICATION
RATS
RODENTS
SEPARATION PROCESSES
SPECTROSCOPY
SYNTHESIS
TUMOR CELLS
VERTEBRATES