Gotoh Y, Sumimoto H, Minakami S
Department of Biochemistry, Kyushu University School of Medicine, Fakuoka, Japan.
Biochim Biophys Acta. 1990 Mar 12;1043(1):52-6. doi: 10.1016/0005-2760(90)90109-b.
When 20-hydroxyleukotriene B4 (20-OH-LTB4) is incubated at pH 10.5 in the presence of NAD+ with an alcohol dehydrogenase isolated from human neutrophils, a polar product is formed as detected on reverse-phase high-performance liquid chromatography (RP-HPLC). The product is identified as 20-oxo-LTB4 (20-CHO-LTB4) on the basis of its co-elution with the authentic compound on HPLC, ultraviolet spectrometry and gas chromatography-mass spectrometry. The 20-CHO-LTB4-forming activity requires NAD+, but NADP+ scarcely replaces NAD+. The apparent Km for 20-OH-LTB4 is 83 microM and the Vmax is 2.04 mumol/min per mg of protein. The activity is inhibited by omega-hydroxy fatty acids such as 12-hydroxylauric acid, 16-hydroxypalmitic acid and 12(S), 20-dihydroxyeicosatetraenoic acid, but not by 4-methylpyrazole. At pH 7.0 with NADH, the purified dehydrogenase catalyzes the reverse reaction, the reduction of 20-CHO-LTB4 to 20-OH-LTB4.
当在pH 10.5条件下，于NAD⁺存在时，将20-羟基白三烯B4（20-OH-LTB4）与人中性粒细胞分离的醇脱氢酶一起孵育，通过反相高效液相色谱（RP-HPLC）检测到会形成一种极性产物。根据该产物在HPLC上与标准品的共洗脱情况、紫外光谱以及气相色谱-质谱分析，该产物被鉴定为20-氧代-LTB4（20-CHO-LTB4）。形成20-CHO-LTB4的活性需要NAD⁺，但NADP⁺几乎不能替代NAD⁺。20-OH-LTB4的表观Km为83微摩尔，Vmax为每毫克蛋白质2.04微摩尔/分钟。该活性受到ω-羟基脂肪酸如12-羟基月桂酸、16-羟基棕榈酸和12(S),20-二羟基二十碳四烯酸的抑制，但不受4-甲基吡唑的抑制。在pH 7.0以及NADH存在的条件下，纯化的脱氢酶催化逆反应，即将20-CHO-LTB4还原为20-OH-LTB4。
