Gardner H W, Hamberg M
National Center for Agricultural Utilization Research, Agricultural Research Service, United States Department of Agriculture, Peoria, Illinois 61604.
J Biol Chem. 1993 Apr 5;268(10):6971-7.
Incubation of (3Z)-nonenal (NON) with the 269,000-g particle fraction of seed homogenate of the broad bean (Vicia faba L.) afforded (2E)-4-hydroxy-2-nonenal (HNE) as the principal product. One pathway of HNE formation consisted of initial oxygenation of NON into (2E)-4-hydroperoxy-2-nonenal (HPNE) by a novel (3Z)-alkenal oxygenase activity, followed by conversion of HPNE into HNE by a previously recognized hydroperoxide-dependent epoxygenase. The hydroperoxide intermediate was detected in coincubations of NON and oleic acid, in which experiments the HPNE generated from NON supported epoxygenase-catalyzed epoxidation of oleic acid into 9,10-epoxystearic acid. Furthermore, by using an enzyme preparation in which the epoxygenase had been inactivated by pretreatment with hydrogen peroxide it was possible to isolate and characterize racemic (4R,4S) HPNE following incubation of NON. Although the (3Z)-alkenal oxygenase resembled a lipoxygenase in its action, it was not inhibited by the lipoxygenase inhibitors, 5,8,11,14-eicosatetraynoic acid and nordihydroguaiaretic acid. In a second pathway, HNE was produced by rearrangement of 3,4-epoxynonenal, which was in turn formed from NON by a reaction catalyzed by hydroperoxide-dependent epoxygenase. Support for this pathway came from experiments in which 18O-labeled HNE was isolated following coincubation of NON and 13-18O-labeled linoleic acid 13-hydroperoxide. The existence of 3,4-epoxynonenal as a transient intermediate in HNE biosynthesis was further demonstrated by the isolation of 3,4-epoxynonenal (61% (4R)-configuration) as a trapping product in short time incubations interrupted by addition of sodium borohydride. The two pathways established for biosynthesis of HNE involved the hydroperoxide-reducing and the olefin-epoxidizing activities of hydroperoxide-dependent epoxygenase. In the absence of extraneous olefins and hydroperoxides the two pathways would be tightly coupled and follow the stoichiometry: 2NON + 1O2-->2HNE. It was also shown that the V. faba particle fraction catalyzed oxygenation of (3Z)-hexenal into (2E)-4-hydroxy-2-hexenal.
将（3Z）-壬烯醛（NON）与蚕豆（Vicia faba L.）种子匀浆的269,000 g颗粒部分一起孵育，得到（2E）-4-羟基-2-壬烯醛（HNE）作为主要产物。HNE形成的一条途径包括通过一种新型的（3Z）-烯醛加氧酶活性将NON初步氧化为（2E）-4-氢过氧-2-壬烯醛（HPNE），随后通过先前公认的氢过氧化物依赖性环氧化酶将HPNE转化为HNE。在NON与油酸的共孵育实验中检测到了氢过氧化物中间体，在这些实验中，由NON生成的HPNE支持环氧化酶催化油酸环氧化为9,10-环氧硬脂酸。此外，通过使用一种环氧化酶已用过氧化氢预处理而失活的酶制剂，在NON孵育后可以分离并表征外消旋（4R,4S）HPNE。尽管（3Z）-烯醛加氧酶在其作用上类似于脂氧合酶，但它不受脂氧合酶抑制剂5,8,11,14-二十碳四烯酸和去甲二氢愈创木酸的抑制。在第二条途径中，HNE由3,4-环氧壬烯重排产生，而3,4-环氧壬烯又是由NON通过氢过氧化物依赖性环氧化酶催化的反应形成的。这一途径的证据来自于NON与13-18O标记的亚油酸13-氢过氧化物共孵育后分离出18O标记的HNE的实验。通过在加入硼氢化钠中断的短时间孵育中分离出3,4-环氧壬烯（61%（4R）构型）作为捕获产物，进一步证明了3,4-环氧壬烯作为HNE生物合成中的瞬态中间体的存在。为HNE生物合成建立的两条途径涉及氢过氧化物依赖性环氧化酶的氢过氧化物还原和烯烃环氧化活性。在没有外源烯烃和氢过氧化物的情况下，这两条途径将紧密偶联并遵循化学计量关系：2NON + 1O2→2HNE。还表明蚕豆颗粒部分催化（3Z）-己烯醛氧化为（2E）-4-羟基-2-己烯醛。
