Swedish Environmental Research Institute, Box 21060, S-100 31 Stockholm, Sweden.
Appl Environ Microbiol. 1988 Feb;54(2):524-30. doi: 10.1128/aem.54.2.524-530.1988.
O-methylation of 2,6-dibromophenol was studied in cell extracts prepared from Rhodococcus sp. strain 1395. O-methylation activity was enhanced by the addition of S-adenosyl-l-methionine but was not affected by the addition of 5-methyltetrahydrofolate nor by up to 10 mM MgCl(2) or EDTA. By using 2,6-dibromophenol, 4,5,6-trichloroguaiacol, and pentachlorothiophenol as the substrates, O-methylation activity was also demonstrated in extracts from two other Rhodococcus sp. strains, an Acinetobacter sp. strain, and a Pseudomonas sp. strain. A diverse range of chloro- and bromophenols, chlorothiophenols, chloro- and bromoguaiacols, and chloro- and bromocatechols were assayed as the substrates by using extracts prepared from strain 1395; all of the compounds were methylated to the corresponding anisoles, veratroles, or guaiacols, which have been identified previously from experiments using whole cells. The specific activity of the enzyme towards the thiophenols was significantly higher than it was towards all the other substrates-high activity was found with pentafluorothiophenol, although the activity with pentafluorophenol was undetectable with the incubation times used. For the chlorophenols, the position of the substituents was of cardinal importance. The enzyme had higher activity towards the halogenated catechols than towards the corresponding guaiacols, and selective O-methylation of the 3,4,5-trihalogenocatechols yielded predominantly the 3,4,5-trihalogenoguaiacols. As in experiments with whole cells, neither 2,4-dinitrophenol, hexachlorophene, nor 5-chloro- or 5-bromovanillin was O-methylated. The results showed conclusively that the methylation reactions were enzymatic and confirmed the conclusion from extensive studies using whole cells that methylation of halogenated phenols may be a significant alternative to biodegradation.
2,6-二溴苯酚的 O-甲基化在 Rhodococcus sp. 1395 菌株的细胞提取物中进行了研究。S-腺苷甲硫氨酸的添加增强了 O-甲基化活性，但 5-甲基四氢叶酸或高达 10 mM 的 MgCl 2 或 EDTA 的添加对其没有影响。使用 2,6-二溴苯酚、4,5,6-三氯愈创木酚和五氯噻吩酚作为底物，也在另外两株 Rhodococcus sp. 菌株、一株不动杆菌属菌株和一株假单胞菌属菌株的提取物中证明了 O-甲基化活性。用 1395 菌株制备的提取物作为底物，对各种氯代和溴代苯酚、氯代和溴代愈创木酚、氯代和溴代邻苯二酚以及氯代和溴代邻苯二酚进行了测定；所有化合物均被甲基化为相应的苯甲醚、藜芦醇或愈创木酚，这些化合物先前已从使用全细胞的实验中得到鉴定。该酶对噻吩酚的比活显著高于其他所有底物，对五氟噻吩酚的活性很高，尽管使用的孵育时间使五氟苯酚的活性无法检测到。对于氯代苯酚，取代基的位置至关重要。该酶对卤代儿茶酚的活性高于相应的愈创木酚，并且 3,4,5-三卤代儿茶酚的选择性 O-甲基化主要生成 3,4,5-三卤代愈创木酚。与全细胞实验一样，2,4-二硝基苯酚、六氯酚和 5-氯或 5-溴藜芦醇均未发生 O-甲基化。结果明确表明，这些甲基化反应是酶促的，并证实了使用全细胞进行的广泛研究的结论，即卤代苯酚的甲基化可能是生物降解的一种重要替代方法。
