Lipoxygenase is an abundant protein in cucumber exudates

The presence of lipoxygenase (LOX) has been reported in many plant organs. High LOX activity (1-2 μkatal/mg protein) was detected in exudates from cut cucumber (Cucumis sativus L.) stems and petioles. Exudate LOX had a pH optimum of 5.0, an estimated molecular weight of 95 kDa and cross-reacted on sodium-dodecyl-sulfate gels with anti-LOX antibodies raised against soybean leaf LOX isoenzymes. Lipoxygenase activity was detected on native gels stained with o-dianisidine using linoleic acid as a substrate. Enzyme activity was similar with linoleic and linolenic acid and 2 times greater with arachidonic acid as substrate. At pH 6.8, LOX metabolized linoleic acid into 13- and 9-hydroperoxides at a ratio of 1:2. Linolenic acid was preferentially oxidized at carbon 13. Lipoxygenase activity was inhibited by n-propyl gallate (IC₅₀ 300 nM) and nordihydroguaiaretic acid (IC₅₀ 25 nM), but not by nonsteroidal anti-inflammatory drugs. LOX activity was enhanced 4.5-fold by 300 mM Ca²⁺. Spermine at 1 mM, and putrescine and spermidine at 2 mM completely inhibited LOX activity, but at low concentrations spermine (100 mM) and spermidine (100-500 mM) significantly stimulated LOX activity: 8- and 4.5-fold, respectively. Tissue printing of stem, petiole and hypocotyl sections with subsequent incubation with the antiserum raised against soybean leaf LOX revealed the presence of LOX in the internal and external phloem and in the sieve tubes.