In this work, the gene of conjugated linoleic acids hydrase (CLA-HY) was cloned from the L. plantarum p-8, and the protein of CLA-HY was expressed in the E. coli BL21. GC-MS was employed to verify that the purified CLA-HY can convert Linoleic Acid (LA) into 10-hydroxy-cis-12-octadecenoic acid (10-HOE) in the presence of FAD. And the optimal pH and temperature for maximizing CLA-HY catalytic activity were discovered as 6.0 and 35 °C, respectively. In addition, the catalytic ability of CLA-HY can be inhibited by plenty of cations such as Mg²⁺, Mn²⁺, Zn²⁺, Cu²⁺, Fe²⁺, Fe³⁺, Ni²⁺ and Ca²⁺. Finally, the K_m, V_max, K_cat and K_cat/K_m of CLA-HY were determined as 7.62 mM, 2.59 mM h⁻¹, 8.33 × 10³ h⁻¹ and 1.09 × 10³ mM⁻¹ h⁻¹, respectively. Moreover, both M76 and G74 residues played significant roles on catalyzing LA to 10-HOE using the site-directed mutation technology and molecular simulations.

中文翻译：

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植物乳杆菌p-8的两个CLA水合突变体的动力学和功能。

在这项工作中，从植物乳杆菌p -8克隆了共轭亚油酸水合酶（CLA-HY）的基因，并在大肠杆菌BL21中表达了CLA-HY的蛋白质。使用GC-MS验证纯化的CLA-HY在FAD存在下能否将亚油酸（LA）转化为10-羟基-顺式-12-十八烯酸（10-HOE）。最大化CLA-HY催化活性的最佳pH和温度分别为6.0和35°C。另外，Mg 2 ⁺，Mn 2 ⁺，Zn 2 ⁺，Cu 2 ⁺，Fe 2 ⁺，Fe ³⁺，Ni ²⁺和Ca ²⁺等大量阳离子可抑制CLA-HY的催化能力。。最后，K个_米，V_最大，K_猫和K_猫/ K_米CLA-HY的确定为7.62毫米，2.59毫米高^-1，8.33×10 ³ ħ ^-1和1.09×10 ³毫米^-1 ħ ^{- 1}。此外，使用定点突变技术和分子模拟，M76和G74残基均在催化LA生成10-HOE方面发挥重要作用。