Rabbit reticulocyte lipoxygenase catalyzes specific 12(S) and 15(S) oxygenation of arachidonoyl-phosphatidylcholine.
Rabbit reticulocyte lipoxygenase catalyzes specific 12(S) and 15(S) oxygenation of arachidonoyl-phosphatidylcholine.
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兔网织红细胞脂氧合酶催化花生四烯酰磷脂酰胆碱的特异性 12(S) 和 15(S) 氧化。
DOI：
10.1016/0003-9861(88)90156-7
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发表时间：
1988
影响因子：
3.9
通讯作者：
Brash,AR
中科院分区：
文献类型：
--
作者：
Murray,JJ;Brash,AR
已知兔网织红细胞脂氧合酶显示出不寻常的酯化多不饱和脂肪酸的氧合设施，但产品的精确结构尚不清楚。以游离花生四烯酸盐作为底物，已知该酶催化15和12 S氧化，并且证明在磷脂上催化这些反应的设施将扩展细胞中脂氧合酶反应的潜在范围。我们选择详细研究酶与天然磷脂，棕榈酰/花生四烯酸-磷脂酰胆碱的反应。我们确定了产品的性质，通过RP-HPLC的初步分离，然后通过HPLC，UV，GC-MS和HPLC的羟基构型的立体分析的含氧产物的酯交换和鉴定。主要产物被鉴定为磷脂酰胆碱，其中花生四烯酸组分转化为15（S）-过氧化氢-二十碳四烯酸。第二种含氧磷脂以较小的量产生（后一种产物的2-5%），并被鉴定为12（S）-含氧类似物。这些产品也被确定后的网织红细胞脂氧合酶与人红细胞膜的反应，其中放射性标记的预孵育与[3 H]花生四烯酸。12 S氧合的发现代表了第一个证据，即脂氧合酶可以控制以花生四烯酸磷脂的10-碳为中心的反应。这是一个重要的先例，因为从碳-10提取氢是脂氧合酶催化合成8-和12-氢过氧二十碳四烯酸（HPETE）以及将5-和15-HPETE转化为白三烯的关键步骤。
The rabbit reticulocyte lipoxygenase is known to display an unusual facility for oxygenation of esterified polyunsaturated fatty acids, yet the precise structures of the products are not known. With free arachidonate as substrate the enzyme is known to catalyze 15Sand 12Soxygenations, and demonstration of a facility for catalysis of these reactions on phospholipids would extend the potential scope of lipoxygenase reactions in cells. We elected to study in detail the reaction of the enzyme with a natural phospholipid, palmitoyl/arachidonoyl-phosphatidylcholine. We determined the nature of the products by initial isolation by RP-HPLC, followed by transesterification and identification of the oxygenated products by HPLC, uv, GC-MS, and steric analysis of hydroxyl configuration by HPLC. The major product was identified as a phosphatidylcholine in which the arachidonate component was converted to the 15(S)-hydroperoxy-eicosatetraenoate. A second oxygenated phospholipid was produced in smaller quantities (2–5% of the latter product) and identified as the 12(S)-oxygenated analog. These products were also identified after reaction of the reticulocyte lipoxygenase with human red cell membranes which were radiolabeled by preincubation with [3H]arachidonic acid. The finding of 12Soxygenation represents the first evidence that a lipoxygenase can control a reaction centered on the 10-carbon of an arachidonoyl phospholipid. This is an important precedent, because hydrogen abstraction from carbon-10 is a critical step in the lipoxygenase-catalyzed synthesis of 8- and 12-hydroperoxyeicosatetraenoates (HPETEs) and for the conversion of 5- and 15-HPETEs to leukotrienes.