Pérez-Pantoja Danilo, Donoso Raúl A, Sánchez Miguel A, González Bernardo
Departamento de Genética Molecular y Microbiología, Facultad de Ciencias Biológicas, P. Universidad Católica de Chile, Millennium Nucleus on Microbial Ecology and Environmental Microbiology and Biotechnology, NM-EMBA, Center for Advanced Studies in Ecology and Biodiversity, CASEB, Santiago, Chile.
Facultad de Ingeniería y Ciencia, Universidad Adolfo Ibáñez, Santiago, Chile.
Microbiology (Reading). 2009 Nov;155(Pt 11):3641-3651. doi: 10.1099/mic.0.032086-0. Epub 2009 Aug 14.
Maleylacetate reductases (MAR) are required for biodegradation of several substituted aromatic compounds. To date, the functionality of two MAR-encoding genes (tfdF(I) and tfdF(II)) has been reported in Cupriavidus necator JMP134(pJP4), a known degrader of aromatic compounds. These two genes are located in tfd gene clusters involved in the turnover of 2,4-dichlorophenoxyacetate (2,4-D) and 3-chlorobenzoate (3-CB). The C. necator JMP134 genome comprises at least three other genes that putatively encode MAR (tcpD, hqoD and hxqD), but confirmation of their functionality and their role in the catabolism of haloaromatic compounds has not been assessed. RT-PCR expression analyses of C. necator JMP134 cells exposed to 2,4-D, 3-CB, 2,4,6-trichlorophenol (2,4,6-TCP) or 4-fluorobenzoate (4-FB) showed that tfdF(I) and tfdF(II) are induced by haloaromatics channelled to halocatechols as intermediates. In contrast, 2,4,6-TCP only induces tcpD, and any haloaromatic compounds tested did not induce hxqD and hqoD. However, the tcpD, hxqD and hqoD gene products showed MAR activity in cell extracts and provided the MAR function for 2,4-D catabolism when heterologously expressed in MAR-lacking strains. Growth tests for mutants of the five MAR-encoding genes in strain JMP134 showed that none of these genes is essential for degradation of the tested compounds. However, the role of tfdF(I)/tfdF(II) and tcpD genes in the expression of MAR activity during catabolism of 2,4-D and 2,4,6-TCP, respectively, was confirmed by enzyme activity tests in mutants. These results reveal a striking example of genetic redundancy in the degradation of aromatic compounds.
马来酰乙酸还原酶（MAR）是几种取代芳香族化合物生物降解所必需的。迄今为止，在已知的芳香族化合物降解菌——食酸铜绿假单胞菌JMP134(pJP4)中，已报道了两个编码MAR的基因（tfdF(I)和tfdF(II)）的功能。这两个基因位于参与2,4-二氯苯氧基乙酸（2,4-D）和3-氯苯甲酸（3-CB）代谢的tfd基因簇中。食酸铜绿假单胞菌JMP134基因组还包含至少其他三个推测编码MAR的基因（tcpD、hqoD和hxqD），但尚未评估它们的功能及其在卤代芳香族化合物分解代谢中的作用。对暴露于2,4-D、3-CB、2,4,6-三氯苯酚（2,4,6-TCP）或4-氟苯甲酸（4-FB）的食酸铜绿假单胞菌JMP134细胞进行的逆转录聚合酶链反应（RT-PCR）表达分析表明，tfdF(I)和tfdF(II)由作为中间体导向卤代儿茶酚的卤代芳香族化合物诱导。相比之下，2,4,6-TCP仅诱导tcpD，并且所测试的任何卤代芳香族化合物均不诱导hxqD和hqoD。然而，tcpD、hxqD和hqoD基因产物在细胞提取物中显示出MAR活性，并且当在缺乏MAR的菌株中异源表达时，为2,4-D分解代谢提供了MAR功能。对JMP134菌株中五个编码MAR的基因的突变体进行的生长测试表明，这些基因中没有一个对于所测试化合物的降解是必需的。然而，通过对突变体进行酶活性测试，证实了tfdF(I)/tfdF(II)和tcpD基因分别在2,4-D和2,4,6-TCP分解代谢过程中MAR活性表达中的作用。这些结果揭示了芳香族化合物降解中基因冗余的一个显著例子。
