Department of Biotechnology, Burdwan University, Golapbag, Burdwan, West Bengal, 713104, India.
Biomed Res Int. 2013;2013:723680. doi: 10.1155/2013/723680. Epub 2013 Sep 8.
Stenotrophomonas maltophilia PM102 (NCBI GenBank Acc. no. JQ797560) is capable of growth on trichloroethylene as the sole carbon source. In this paper, we report the purification and characterisation of oxygenase present in the PM102 isolate. Enzyme activity was found to be induced 10.3-fold in presence of 0.7 mM copper with a further increment to 14.96-fold in presence of 0.05 mM NADH. Optimum temperature for oxygenase activity was recorded at 36°C. The reported enzyme was found to have enhanced activity at pH 5 and pH 8, indicating presence of two isoforms. Maximum activity was seen on incubation with benzene compared to other substrates like TCE, chloroform, toluene, hexane, and petroleum benzene. K(m) and V(max) for benzene were 3.8 mM and 340 U/mg/min and those for TCE were 2.1 mM and 170 U/mg/min. The crude enzyme was partially purified by ammonium sulphate precipitation followed by dialysis. Zymogram analysis revealed two isoforms in the 70% purified enzyme fraction. The activity stain was more prominent when the native gel was incubated in benzene as substrate in comparison to TCE. Crude enzyme and purified enzyme fractions were assayed for TCE degradation by the Fujiwara test. TCE biotransformation products were analysed by FT-IR spectroscopy.
嗜麦芽寡养单胞菌 PM102（NCBI GenBank 登录号 JQ797560）能够以三氯乙烯为唯一碳源生长。在本文中，我们报告了 PM102 分离物中存在的加氧酶的纯化和特性。在存在 0.7 mM 铜的情况下，酶活性诱导了 10.3 倍，在存在 0.05 mM NADH 的情况下进一步增加到 14.96 倍。氧合酶活性的最适温度记录在 36°C。报道的酶在 pH 5 和 pH 8 下表现出增强的活性，表明存在两种同工酶。与其他底物如 TCE、氯仿、甲苯、己烷和石油苯相比，与苯孵育时酶的活性最高。苯的 K(m)和 V(max)分别为 3.8 mM 和 340 U/mg/min，TCE 的 K(m)和 V(max)分别为 2.1 mM 和 170 U/mg/min。粗酶通过硫酸铵沉淀和透析进行部分纯化。70%纯化酶级分的同工酶分析显示出两种同工酶。与 TCE 相比，当在苯作为底物孵育时，天然凝胶的活性染色更为明显。用 Fujiwara 试验测定粗酶和纯化酶级分对 TCE 的降解。用傅里叶变换红外光谱法分析 TCE 生物转化产物。
