In vitro oxygenation of soybean biomembranes by lipoxygenase-2.
The ability of soybean lipoxygenases-1 and -2 to oxygenate biomembranes isolated from soybean seedlings has been investigated. Constituents of the lipid bilayer were analyzed by means of reversed phase and chiral phase high performance liquid chromatography, gas chromatography/mass spectrometry, high performance thin layer chromatography and uv spectroscopy. Evidence is presented that soybean lipoxygenase-2, at variance with the type-1 enzyme, oxygenates the esterified unsaturated fatty acid moieties in biomembranes, whereas membrane-embedded free unsaturated fatty acid moieties were not a suitable substrate for either isoenzyme. The oxygenation products derived from the biomembranes were the 9- and 13-hydroperoxides of linoleic acid residues, in a molar ratio of 1.0 to 1.7, and the 9- and 13-hydroperoxides of alpha-linolenic acid residues, in a molar ratio of 1.0 to 0.1. The R/S ratios of 13-hydroperoxy-9Z,11E-octadecadienoic acid and 9-hydroperoxy-10E,12Z,15Z-octadecatrienoic acid were found to be 0.5 and 25.0, respectively. These stereospecificity values were much higher than those of hydroperoxides isolated after incubation of lipoxygenase-2 with non-membraneous fatty acids or their methyl esters. The hydroperoxy fatty acids produced were distributed in neutral lipids and phospholipids isolated from soybean membranes, the former being oxidized to a larger extent. Furthermore, both intracellular and plasma membranes were substrates for the enzymic oxygenation, with a preference for those of chloroplasts followed by those of Golgi apparatus, endoplasmic reticulum, plasma membrane and mitochondria. These data point towards a different action of the two lipoxygenases in soybean cells. We suggest that the type-2 enzyme plays a role in the in vivo remodelling of biomembranes. The physiological relevance of these findings is discussed.
Maccarrone
M
M
Bijvoet Center for Biomolecular Research, Department of Bio-organic Chemistry, Utrecht University, The Netherlands.
van Aarle
P G
PG
Veldink
G A
GA
Vliegenthart
J F
JF
D008051:Lipid Bilayers / Q000737:chemistry*; D008084:Lipoxygenase / Q000302:isolation & purification / Q000378:metabolism*; D010100:Oxygen / Q000032:analysis*; D013025:Glycine max / Q000737:chemistry / Q000201:enzymology* / Q000648:ultrastructure; D013347:Subcellular Fractions / Q000737:chemistry
D008051:Lipid Bilayers; D008084:Lipoxygenase; D010100:Oxygen