Perkins E J, Gordon M P, Caceres O, Lurquin P F
Department of Biochemistry, University of Washington, Seattle 98195.
J Bacteriol. 1990 May;172(5):2351-9. doi: 10.1128/jb.172.5.2351-2359.1990.
Growth of Alcaligenes eutrophus JMP134 on 2,4-dichlorophenoxyacetate requires a 2,4-dichlorphenol hydroxylase encoded by gene tfdB. Catabolism of either 2,4-dichlorophenoxyacetate or 3-chlorobenzoate involves enzymes encoded by the chlorocatechol oxidative operon consisting of tfdCDEF, which converts 3-chloro- and 3,5-dichlorocatechol to maleylacetate and chloromaleylacetate, respectively. Transposon mutagenesis has localized tfdB and tfdCDEF to EcoRI fragment B of plasmid pJP4 (R. H. Don, A. J. Wieghtman, H.-J. Knackmuss, and K. N. Timmis, J. Bacteriol. 161:85-90, 1985). We present the complete nucleotide sequence of tfdB and tfdCDEF contained within a 7,954-base-pair HindIII-SstI fragment from EcoRI fragment B. Sequence and expression analysis of tfdB in Escherichia coli suggested that 2,4-dichlorophenol hydroxylase consists of a single subunit of 65 kilodaltons. The amino acid sequences of proteins encoded by tfdD and tfdE were found to be 63 and 53% identical to those of functionally similar enzymes encoded by clcB and clcD, respectively, from plasmid pAC27 of Pseudomonas putida. P. putida(pAC27) can utilize 3-chlorocatechol but not dichlorinated catechols. A region of DNA adjacent to clcD in pAC27 was found to be 47% identical in amino acid sequence to tfdF, a gene important in catabolizing dichlorocatechols. The region in pAC27 does not appear to encode a protein, suggesting that the absence of a functional trans-chlorodienelactone isomerase may prevent P. putida(pAC27) from utilizing 3,5-dichlorocatechol.
嗜麦芽窄食单胞菌JMP134在2,4-二氯苯氧基乙酸上生长需要由基因tfdB编码的2,4-二氯苯酚羟化酶。2,4-二氯苯氧基乙酸或3-氯苯甲酸的分解代谢涉及由由tfdCDEF组成的氯儿茶酚氧化操纵子编码的酶，该操纵子分别将3-氯儿茶酚和3,5-二氯儿茶酚转化为马来酰乙酸和氯马来酰乙酸。转座子诱变已将tfdB和tfdCDEF定位到质粒pJP4的EcoRI片段B上（R. H. 唐、A. J. 维特曼、H.-J. 克纳克穆斯和K. N. 蒂米斯，《细菌学杂志》161:85-90, 1985）。我们给出了来自EcoRI片段B的一个7954碱基对的HindIII-SstI片段中包含的tfdB和tfdCDEF的完整核苷酸序列。tfdB在大肠杆菌中的序列和表达分析表明，2,4-二氯苯酚羟化酶由一个65千道尔顿的单亚基组成。发现由tfdD和tfdE编码的蛋白质的氨基酸序列分别与恶臭假单胞菌质粒pAC27的clcB和clcD编码的功能相似酶的氨基酸序列有63%和53%的同一性。恶臭假单胞菌（pAC27）可以利用3-氯儿茶酚，但不能利用二氯儿茶酚。发现pAC27中与clcD相邻的一个DNA区域在氨基酸序列上与tfdF有47%的同一性，tfdF是一种在二氯儿茶酚分解代谢中起重要作用的基因。pAC27中的该区域似乎不编码一种蛋白质，这表明缺乏功能性的反式氯二烯内酯异构酶可能阻止恶臭假单胞菌（pAC27）利用3,5-二氯儿茶酚。
