Department of Microbial Pathogenesis, Helmholtz Zentrum für Infektionsforschung, Braunschweig, Germany.
J Bacteriol. 2010 Mar;192(6):1543-52. doi: 10.1128/JB.01208-09. Epub 2010 Jan 8.
Catechols are central intermediates in the metabolism of aromatic compounds. Degradation of 4-methylcatechol via intradiol cleavage usually leads to the formation of 4-methylmuconolactone (4-ML) as a dead-end metabolite. Only a few microorganisms are known to mineralize 4-ML. The mml gene cluster of Pseudomonas reinekei MT1, which encodes enzymes involved in the metabolism of 4-ML, is shown here to encode 10 genes found in a 9.4-kb chromosomal region. Reverse transcription assays revealed that these genes form a single operon, where their expression is controlled by two promoters. Promoter fusion assays identified 4-methyl-3-oxoadipate as an inducer. Mineralization of 4-ML is initiated by the 4-methylmuconolactone methylisomerase encoded by mmlI. This reaction produces 3-ML and is followed by a rearrangement of the double bond catalyzed by the methylmuconolactone isomerase encoded by mmlJ. Deletion of mmlL, encoding a protein of the metallo-beta-lactamase superfamily, resulted in a loss of the capability of the strain MT1 to open the lactone ring, suggesting its function as a 4-methyl-3-oxoadipate enol-lactone hydrolase. Further metabolism can be assumed to occur by analogy with reactions known from the 3-oxoadipate pathway. mmlF and mmlG probably encode a 4-methyl-3-oxoadipyl-coenzyme A (CoA) transferase, and the mmlC gene product functions as a thiolase, transforming 4-methyl-3-oxoadipyl-CoA into methylsuccinyl-CoA and acetyl-CoA, as indicated by the accumulation of 4-methyl-3-oxoadipate in the respective deletion mutant. Accumulation of methylsuccinate by an mmlK deletion mutant indicates that the encoded acetyl-CoA hydrolase/transferase is crucial for channeling methylsuccinate into the central metabolism.
儿茶酚是芳香族化合物代谢的重要中间产物。4-甲基儿茶酚通过邻二醇裂解途径通常会导致 4-甲基粘康酸内酯（4-ML）的形成，这是一种终末代谢产物。只有少数微生物能够矿化 4-ML。本文展示了假单胞菌属雷氏不动杆菌 MT1 的 mml 基因簇，该基因簇编码参与 4-ML 代谢的酶，该基因簇位于 9.4kb 的染色体区域内，共编码 10 个基因。逆转录试验表明，这些基因构成一个单一的操纵子，其表达受两个启动子调控。启动子融合试验鉴定出 4-甲基-3-氧代戊二酸为诱导物。4-ML 的矿化是由 mmlI 编码的 4-甲基粘康酸内酯甲基异构酶起始的。该反应产生 3-ML，随后由 mmlJ 编码的甲基粘康酸内酯异构酶催化双键重排。mmlL 编码金属β-内酰胺酶超家族的蛋白，该基因缺失会导致 MT1 丧失开环能力，表明其功能可能是 4-甲基-3-氧代戊二酸烯醇内酯水解酶。进一步的代谢推测可以通过与 3-氧代戊二酸途径已知的反应进行类比。mmlF 和 mmlG 可能编码 4-甲基-3-氧代戊二酰辅酶 A（CoA）转移酶，mmlC 基因产物作为硫解酶，将 4-甲基-3-氧代戊二酰-CoA 转化为甲基琥珀酰-CoA 和乙酰-CoA，这一点可以从相应的缺失突变体中 4-甲基-3-氧代戊二酸的积累得到证明。mmlK 缺失突变体中甲基琥珀酸的积累表明，编码的乙酰-CoA 水解酶/转移酶对于将甲基琥珀酸定向进入中心代谢途径至关重要。
