Gargouri M, Legoy M D
Laboratoire de Génie Protéique et Cellulaire, Pôle Sciences et Technologie, Université de La Rochelle, France.
Biochim Biophys Acta. 1997 Feb 8;1337(2):227-32. doi: 10.1016/s0167-4838(96)00169-0.
Analysis of the kinetic behaviour of a two-enzyme-system carrying out two consecutive reactions was investigated in macroheterogeneous biphasic media (octane/buffer pH 9.6, v/v = 1:1). The lipase-catalysed hydrolysis of trilinolein and the subsequent lipoxygenation of the liberated linoleic acid, were coupled in a modified Lewis cell with a well-defined liquid/liquid interfacial area. Trilinolein was dissolved in the organic phase and hydrolysed in the presence of Mucor javanicus lipase at the organic/aqueous interface. Linoleic acid, liberated after hydrolysis was transferred to the aqueous phase and reacted with lipoxygenase. This reaction consumed linoleic acid and produced hydroperoxides, which favoured the transfer of residual linoleic acid, since they possess surface active properties. Catalysis and transfer influenced each other reciprocally. At low substrate concentrations, cooperativity phenomena were observed in the experimental and also the modelled two-enzyme systems. When the initial substrate concentration was high, the kinetic behaviour of the two-enzyme system in a compartmentalised medium, seemed to be independent of the substrate concentration, unlike that observed in homogeneous monophasic enzymology. The numerical integration program used to model the two-enzyme system was based on results obtained in separate studies of the following three phenomena: (1) trilinolein hydrolysis in biphasic medium. (2) linoleic acid transfer across a liquid/liquid interface and (3) lipoxygenation in an aqueous media. Results obtained by modelling were similar to the results observed experimentally.
在宏观非均相双相介质（辛烷/缓冲液pH 9.6，v/v = 1:1）中研究了进行两个连续反应的双酶系统的动力学行为。三油酸甘油酯的脂肪酶催化水解以及随后释放的亚油酸的脂氧合反应，在具有明确液/液界面面积的改良Lewis池中偶联。三油酸甘油酯溶解在有机相中，并在爪哇毛霉脂肪酶存在下于有机/水界面处水解。水解后释放的亚油酸转移到水相中并与脂氧合酶反应。该反应消耗亚油酸并产生氢过氧化物，由于它们具有表面活性性质，这有利于残留亚油酸的转移。催化和转移相互影响。在低底物浓度下，在实验和模拟的双酶系统中均观察到协同现象。当初始底物浓度较高时，与在均相单相酶学中观察到的情况不同，双酶系统在分隔介质中的动力学行为似乎与底物浓度无关。用于模拟双酶系统的数值积分程序基于以下三个现象的单独研究结果：（1）双相介质中三油酸甘油酯的水解。（2）亚油酸跨液/液界面的转移和（3）水相中脂氧合反应。模拟得到的结果与实验观察结果相似。
