The homogentisate pathway:: A central catabolic pathway involved in the degradation of L-phenylalanine, L-tyrosine, and 3-hydroxyphenylacetate in Pseudomonas putida
The homogentisate pathway:: A central catabolic pathway involved in the degradation of L-phenylalanine, L-tyrosine, and 3-hydroxyphenylacetate in Pseudomonas putida
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DOI：
10.1128/jb.186.15.5062-5077.2004
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发表时间：
2004-08-01
影响因子：
3.2
通讯作者：
Miñambres, B
中科院分区：
文献类型：
--
作者：
Arias-Barrau, E;Olivera, ER;Miñambres, B
恶臭假单胞菌通过外周途径代谢Phe和Tyr，所述外周途径涉及Phe羟基化为Tyr（PhhAB）、Tyr转化为4-羟基苯丙酮酸（TyrB）以及作为中心中间体的尿黑酸（Hpd）的形成。然后通过涉及三种酶的中心分解代谢途径，尿黑酸双加氧酶（HmgA）、富马酰乙酰乙酸水解酶（HmgB）和马来酰乙酰乙酸异构酶（HmgC）分解代谢尿黑酸，最终产生富马酸和乙酰乙酸。而phh、tyr和hpd基因在恶臭假单胞菌基因组中不连锁，hmgABC基因似乎形成单个转录单位。凝胶阻滞试验和lacZ翻译融合实验表明，hmgR编码一个特定的阻遏物，控制诱导表达的分歧转录hmgABC分解代谢基因，和尿黑酸是诱导分子。足迹分析表明，HmgR保护的Phmg启动子中的一个区域，跨越一个17-bp的回文基序和外部直接重复从位置-16到位置29相对于转录起始位点。因此，HmGR蛋白是第一个作为芳香族分解代谢途径的阻遏物的IcIR型调节剂。我们设计了一个广泛的宿主范围可动员的分解代谢盒，包含hmgABC，hpd和tyrB基因，允许异源细菌使用Tyr作为独特的碳源和能源。值得注意的是，我们在这里表明，3-羟基苯乙酸在恶臭假单胞菌U漏斗也进入尿黑酸中央途径，揭示了hmg簇是一个关键的分解代谢性状的少数芳香族化合物的生物降解。
Pseudomonas putida metabolizes Phe and Tyr through a peripheral pathway involving hydroxylation of Phe to Tyr (PhhAB), conversion of Tyr into 4-hydroxyphenylpyruvate (TyrB), and formation of homogentisate (Hpd) as the central intermediate. Homogentisate is then catabolized by a central catabolic pathway that involves three enzymes, homogentisate dioxygenase (HmgA), fumarylacetoacetate hydrolase (HmgB), and maleylacetoacetate isomerase (HmgC), finally yielding fumarate and acetoacetate. Whereas the phh, tyr, and hpd genes are not linked in the P. putida genome, the hmgABC genes appear to form a single transcriptional unit. Gel retardation assays and lacZ translational fusion experiments have shown that hmgR encodes a specific repressor that controls the inducible expression of the divergently transcribed hmgABC catabolic genes, and homogentisate is the inducer molecule. Footprinting analysis revealed that HmgR protects a region in the Phmg promoter that spans a 17-bp palindromic motif and an external direct repetition from position -16 to position 29 with respect to the transcription start site. The HmgR protein is thus the first IcIR-type regulator that acts as a repressor of an aromatic catabolic pathway. We engineered a broad-host-range mobilizable catabolic cassette harboring the hmgABC, hpd, and tyrB genes that allows heterologous bacteria to use Tyr as a unique carbon and energy source. Remarkably, we show here that the catabolism of 3-hydroxyphenylacetate in P. putida U funnels also into the homogentisate central pathway, revealing that the hmg cluster is a key catabolic trait for biodegradation of a small number of aromatic compounds.