Gardner HW, Grove MJ
Bioactive Research, National Center for Agricultural Utilization Research, Agricultural Research Service, United States Department of Agriculture, Peoria, Illinois 61604, USA.
Plant Physiol. 1998 Apr;116(4):1359-66.
In previous work with soybean (Glycine max), it was reported that the initial product of 3Z-nonenal (NON) oxidation is 4-hydroperoxy-2E-nonenal (4-HPNE). 4-HPNE can be converted to 4-hydroxy-2E-nonenal by a hydroperoxide-dependent peroxygenase. In the present work we have attempted to purify the 4-HPNE-producing oxygenase from soybean seed. Chromatography on various supports had shown that O2 uptake with NON substrate consistently coincided with lipoxygenase (LOX)-1 activity. Compared with oxidation of LOX's preferred substrate, linoleic acid, the activity with NON was about 400- to 1000-fold less. Rather than obtaining the expected 4-HPNE, 4-oxo-2E-nonenal was the principal product of NON oxidation, presumably arising from the enzyme-generated alkoxyl radical of 4-HPNE. In further work a precipitous drop in activity was noted upon dilution of LOX-1 concentration; however, activity could be enhanced by spiking the reaction with 13S-hydroperoxy-9Z, 11E-octadecadienoic acid. Under these conditions the principal product of NON oxidation shifted to the expected 4-HPNE. 4-HPNE was demonstrated to be 83% of the 4S-hydroperoxy-stereoisomer. Therefore, LOX-1 is also a 3Z-alkenal oxygenase, and it exerts the same stereospecificity of oxidation as it does with polyunsaturated fatty acids. Two other LOX isozymes of soybean seed were also found to oxidize NON to 4-HPNE with an excess of 4S-hydroperoxy-stereoisomer.
在之前对大豆（Glycine max）的研究中，有报道称3Z-壬烯醛（NON）氧化的初始产物是4-氢过氧-2E-壬烯醛（4-HPNE）。4-HPNE可通过一种依赖氢过氧化物的过氧合酶转化为4-羟基-2E-壬烯醛。在本研究中，我们试图从大豆种子中纯化产生4-HPNE的加氧酶。在各种支持物上进行色谱分析表明，以NON为底物时的氧气摄取始终与脂氧合酶（LOX）-1的活性一致。与LOX的首选底物亚油酸的氧化相比，NON的活性约低400至1000倍。NON氧化的主要产物不是预期的4-HPNE，而是4-氧代-2E-壬烯醛，推测是由酶产生的4-HPNE烷氧基自由基产生的。在进一步的研究中，发现稀释LOX-1浓度后活性急剧下降；然而，通过在反应中加入13S-氢过氧-9Z，11E-十八碳二烯酸可以提高活性。在这些条件下，NON氧化的主要产物转变为预期的4-HPNE。已证明4-HPNE占4S-氢过氧立体异构体的83%。因此，LOX-1也是一种3Z-烯醛加氧酶，并且它对氧化具有与多不饱和脂肪酸相同的立体特异性。还发现大豆种子的另外两种LOX同工酶也将NON氧化为4-HPNE，且4S-氢过氧立体异构体过量。
