title: Enzyme with dual lipoxygenase activities catalyzes leukotriene A4 synthesis from arachidonic acid.
abstract: When arachidonic acid was incubated with homogenates of potato tubers, two isomers of 6-trans-leukotriene B4, epimeric at C-12, were formed in addition to the major product, (5S-hydroperoxy-6-trans-8,11,14-cis-icosatetraenoic acid (5-HPETE). To elucidate the mechanism of biosynthesis of the dihydroxy-acids, the lipoxygenase from the potato tubers was purified to apparent homogeneity by a combination of conventional chromatographic procedures and high-performance liquid chromatography equipped with a chromatofocusing column (Mono-P). The purified lipoxygenase acted on arachidonic acid and bishomo-gamma-linolenic acid to yield (5S)-hydroperoxy- and (8S)-hydroperoxyicosanoids, respectively. Furthermore, the purified enzyme converted 5-HPETE to leukotriene A4, with the presence of the epoxide intermediate being demonstrated by 18O2 experiments, methanol trapping, as well as further conversion to leukotriene B4 by the purified leukotriene A4 hydrolase. Several experiments, including those with lipoxygenase inhibitors, heat treatment, and competitive inhibition, indicated that both the 5-lipoxygenase and leukotriene A4 synthase activities resided in the same protein and that the formation of leukotriene A4 from 5-HPETE was catalyzed by the 8-lipoxygenase activity of the enzyme.
mesh_terms: D016718:Arachidonic Acid; D001095:Arachidonic Acids / Q000096:biosynthesis* / Q000378:metabolism*; D002250:Carbon Radioisotopes; D007700:Kinetics; D017572:Leukotriene A4; D008084:Lipoxygenase / Q000302:isolation & purification / Q000378:metabolism*; D010944:Plants / Q000201:enzymology
chemical_list: D001095:Arachidonic Acids; D002250:Carbon Radioisotopes; D017572:Leukotriene A4; D016718:Arachidonic Acid; D008084:Lipoxygenase