Matsuda Y, Beppu T, Arima K
Biochim Biophys Acta. 1978 Sep 28;530(3):439-50.
A lipoxygenase obtained from the fungus Fusarium oxysporum was purified and crystallized. Using the purified enzyme, the positional specificity of linoleate peroxidation was studied. Linoleate hydroperoxides were converted into the corresponding trimethylsilyl derivative by reduction, catalytic hydrogenation and treatment with hexamethyldisilazane/trimethylchlorosilane/pyridine and then analyzed by combined gas-liquid chromatography-mass spectrometry. Fusarium lipoxygenase was found to produce 9- or 13-hydroperoxy-octadecadienoates from linoleate. The ratio of 9- to 13-hydroperoxides produced by the enzyme was also determined by high performance liquid chromatography of their methyl esters. When the enzymic reaction proceeded at pH 9.0 and 12.0, the ratio of 9- to 13-hydroperoxide isomers was 70 : 30 and 56 : 44, respectively. With the use of the heavy isotope of oxygen (18O2), atoms of oxygen introduced into hydroperoxides were found to be derived from the gaseous phase and not from the aqueous phase.
从尖孢镰刀菌中获得的一种脂氧合酶被纯化并结晶。使用纯化后的酶，研究了亚油酸过氧化反应的位置特异性。亚油酸氢过氧化物通过还原、催化氢化以及用六甲基二硅氮烷/三甲基氯硅烷/吡啶处理，转化为相应的三甲基硅烷基衍生物，然后通过气液色谱 - 质谱联用进行分析。发现镰刀菌脂氧合酶能从亚油酸生成9 - 或13 - 氢过氧十八碳二烯酸酯。该酶产生的9 - 与13 - 氢过氧化物的比例也通过其甲酯的高效液相色谱法测定。当酶促反应在pH 9.0和12.0下进行时，9 - 与13 - 氢过氧化物异构体的比例分别为70 : 30和56 : 44。通过使用氧的重同位素（(^{18}O_2)），发现引入到氢过氧化物中的氧原子来自气相而非水相。
