Molecular characterization of the Entner-Doudoroff pathway in Escherichia coli: sequence analysis and localization of promoters for the edd-eda operon

Egan, Suhelen; Fliege, Ronda; Tong, Suxiang; Shibata, Annemarie; Wolf, Richard M.; Conway, Tyrrell

doi:10.1128/jb.174.14.4638-4646.1992

Cited by 83 publications

(78 citation statements)

References 39 publications

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“…The amino acid sequence predicted by the ORF exhibited 60% identity with and 76% similarity to the amino acid sequence of EDD from E. coli (21) and 55% identity with and 71% similarity to that of EDD from Z. mobilis (3) (Fig. 4).…”

Section: Resultsmentioning

confidence: 99%

“…Computer analysis of initial sequence data revealed an ORF extending from within the 247-bp fragment in pPZ196 whose predicted amino acid sequence exhibited a high degree of homology with the EDD sequences from E. coli (21) and Zymomonas mobilis (3). The complete edd sequence from P. aeruginosa consisted of 1,833 bp encoding a 610-amino-acid protein with a predicted molecular weight of 65,140 (Fig.…”

Section: Resultsmentioning

confidence: 99%

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Two genes for carbohydrate catabolism are divergently transcribed from a region of DNA containing the hexC locus in Pseudomonas aeruginosa PAO1

et al. 1994

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The hexC locus ofPseudomonas aeruginosa PAO1 was localized to a 247-bp segment of chromosomal DNA on the multicopy broad-host-range vector pRO1614. The presence of this plasmid (pPZ196) in strain PAO1 produced the so-called "hexC effect," a two-to ninefold increase in the activities of four carbohydrate catabolism enzymes, glucokinase, glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydratase, and 2-keto-3-deoxy-6-phosphogluconate aldolase. The extent of the hexC effect was restricted, since three independently regulated metabolic enzymes were not affected by the presence of the hexC plasmid. Furthermore, the hexC-containing plasmid did not suppress catabolite repression control. Nucleotide sequence analysis of the segment of DNA encompassing hexC revealed a 128-bp region rich in adenosine-plus-thymine (AT) content separating two divergent open reading frames (ORFs). Transcriptional start sites for these two genes were mapped to the intergenic region, demonstrating that this sequence contained overlapping divergent promoters. The intergenic region contained potential regulatory sequences such as dyad symmetry motifs, polydeoxyadenosine tracts, and a sequence matching the integration host factor recognition site in Escherichia coli. One of the ORFs encoded a 610-amino-acid protein with 55 to 60%Yo identity to 6-phosphogluconate dehydratase from E. coli and Zymomonas mobiis. The second ORF coded for a protein of 335 amino acids that displayed 45 to 60% identity to the NAD-dependent glyceraldehyde-3-phosphate dehydrogenase (GAP) family of enzymes. The NAD-dependent GAP gene on the P. aeruginosa chromosome was previously unmapped. GAP was found to exhibit the hexC-dependent increase in its basal activity, establishing it as a fifth catabolic enzyme in the multioperonic hex regulon.Pseudomonads utilize a number of alternative pathways for the catabolism of carbohydrates to carbon and energy (reviewed in reference 37). In Pseudomonas aeruginosa, the coordinately regulated enzymes glucokinase (GLK), glucose-6-phosphate dehydrogenase (ZWF), and the Entner-Doudoroff pathway enzymes 6-phosphogluconate dehydratase (EDD) and 2-keto-3-deoxy-6-phosphogluconate aldolase (EDA) play a central role in the dissimilation of glucose to pyruvate and glyceraldehyde-3-phosphate (Fig. 1). These enzymes are coinduced by growth of P. aeruginosa on such diverse carbohydrates as glucose, gluconate, mannitol, and glycerol (33, 37).Also, their expression is subject to catabolite repression control (CRC) by succinate or other intermediates of the tricarboxylic acid (TCA) cycle (40,58).The genes encoding these four enzymes (glk, zwf, edd, and eda) were mapped by transductional analysis to 39 min on the PAO1 chromosome (14,30,47). Cloning and complementation analyses revealed, however, that the four genes were expressed from separate promoters (15,57), indicating that the genes were not contained in a single operon. Two regulatory effects further defined this group of genes. The first was the hexR mutation, thus far inseparable from edd,...

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Two genes for carbohydrate catabolism are divergently transcribed from a region of DNA containing the hexC locus in Pseudomonas aeruginosa PAO1

et al. 1994

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“…Whereas the D-arabinonate dehydratase from S. solfataricus belongs to COG4948, the same function seems to be performed by members of COG0129 that are commonly annotated as dihydroxyacid dehydratases (IlvD) or 6-phosphogluconate dehydratases (Edd) (67). A member of this family has recently been characterized from S. solfataricus (DHAD, Sso3107), which revealed a broad substrate specificity for aldonic acids (68).…”

Section: Resultsmentioning

confidence: 99%

Identification of the Missing Links in Prokaryotic Pentose Oxidation Pathways

Brouns

Walther

Snijders

et al. 2006

Journal of Biological Chemistry

106

188

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The pentose metabolism of Archaea is largely unknown. Here, we have employed an integrated genomics approach including DNA microarray and proteomics analyses to elucidate the catabolic pathway for D-arabinose in Sulfolobus solfataricus. During growth on this sugar, a small set of genes appeared to be differentially expressed compared with growth on D-glucose. These genes were heterologously overexpressed in Escherichia coli, and the recombinant proteins were purified and biochemically studied. This showed that D-arabinose is oxidized to 2-oxoglutarate by the consecutive action of a number of previously uncharacterized enzymes, including a D-arabinose dehydrogenase, a D-arabinonate dehydratase, a novel 2-keto-3-deoxy-D-arabinonate dehydratase, and a 2,5-dioxopentanoate dehydrogenase. Promoter analysis of these genes revealed a palindromic sequence upstream of the TATA box, which is likely to be involved in their concerted transcriptional control. Integration of the obtained biochemical data with genomic context analysis strongly suggests the occurrence of pentose oxidation pathways in both Archaea and Bacteria, and predicts the involvement of additional enzyme components. Moreover, it revealed striking genetic similarities between the catabolic pathways for pentoses, hexaric acids, and hydroxyproline degradation, which support the theory of metabolic pathway genesis by enzyme recruitment.Pentose sugars are a ubiquitous class of carbohydrates with diverse biological functions. Ribose and deoxyribose are major constituents of nucleic acids, whereas arabinose and xylose are building blocks of several plant cell wall polysaccharides. Many prokaryotes, as well as yeasts and fungi, are able to degrade these polysaccharides, and use the released five-carbon sugars as a sole carbon and energy source. At present, three main catabolic pathways have been described for pentoses. The first is present in Bacteria and uses isomerases, kinases, and epimerases to convert D-and L-arabinose (Ara) and D-xylose (Xyl) into D-xylulose 5-phosphate (Fig. 1A), which is further metabolized by the enzymes of the phosphoketolase or pentose phosphate pathway. The genes encoding the pentose-converting enzymes are often located in gene clusters in bacterial genomes, for example, the araBAD operon for L-Ara (1), the xylAB operon for D-Xyl (2), and the darK-fucPIK gene cluster for D-Ara (3). The second catabolic pathway for pentoses converts D-Xyl into D-xylulose 5-phosphate as well, but the conversions are catalyzed by reductases and dehydrogenases instead of isomerases and epimerases (Fig. 1B). This pathway is commonly found in yeasts, fungi, mammals, and plants, but also in some bacteria (4 -6). In a third pathway, pentoses such as L-Ara, D-Xyl, D-ribose, and D-Ara are metabolized non-phosphorylatively to either 2-oxoglutarate (2-OG) 4 or to pyruvate and glycolaldehyde (Fig. 1C). The conversion to 2-OG, which is a tricarboxylic acid cycle intermediate, proceeds via the subsequent action of a pentose dehydrogenase, a pentonolactonase, a pentoni...

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“…Although this reaction has not been shown to occur in vivo, cleavage of KHG could serve in tricarboxylic acid (TCA) cycle regulation or, when operating in the reverse direction, in the detoxification of glyoxylate. Cloning and sequencing the genes encoding these functions revealed that KHG aldolase and Eda are the same enzyme (7,18). However, this study is the first to extensively characterize eda regulation.…”

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confidence: 98%

Multiple Regulators Control Expression of the Entner-Doudoroff Aldolase (Eda) of Escherichia coli

Murray

Conway

2005

J Bacteriol

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The Escherichia coli eda gene, which encodes the Entner-Doudoroff aldolase, is central to the catabolism of several sugar acids. Here, we show that Eda synthesis is induced by growth on gluconate, glucuronate, or methyl-␤-D-glucuronide; phosphate limitation; and carbon starvation. Transcription of eda initiates from three promoters, designated P1, P2, and P4, each of which is responsible for induction under different growth conditions. P1 controls eda induction on gluconate and is regulated by GntR. P2 controls eda induction on glucuronate and galacturonate and is regulated by KdgR. P4 is active under conditions of phosphate starvation and is directly controlled by PhoB. In addition, CsrA activates Eda synthesis, apparently by an indirect mechanism that may be involved in the modest changes in expression level that are associated with carbon starvation. The complex regulation of eda is discussed with respect to its several physiological roles, which apparently accommodate not only sugar acid catabolism but also detoxification of metabolites that could accumulate during starvation-induced stress.Sugar acids are metabolized via the Entner-Doudoroff and Ashwell pathways (1,5,8,20). The Entner-Doudoroff aldolase, encoded by the eda gene, is necessary for growth on several mucus-derived sugar acids (20), which may define a niche for Escherichia coli colonization of the mammalian large intestine (4). Mutants lacking the key enzyme of the Entner-Doudoroff pathway, 6-phosphogluconate dehydratase (encoded by edd), are compromised in their ability to colonize the mouse intestine (4), as are eda mutants (29).Eda cleavage of 2-keto-3-deoxy-6-phosphogluconate (KDPG) to pyruvate and triose-3-phosphate is a common step of hexonate and hexuronate catabolism (20). Eda has also been found to have a role in the degradation of 2-keto-4-hydroxyglutarate (KHG) to pyruvate and glyoxylate (16). Although this reaction has not been shown to occur in vivo, cleavage of KHG could serve in tricarboxylic acid (TCA) cycle regulation or, when operating in the reverse direction, in the detoxification of glyoxylate. Cloning and sequencing the genes encoding these functions revealed that KHG aldolase and Eda are the same enzyme (7, 18). However, this study is the first to extensively characterize eda regulation.The eda gene is located immediately downstream of edd, at 40.5 min on the E. coli chromosome (Fig. 1) (6). Previously, primer extension analysis of the eda regulatory region revealed four putative eda promoters (7). P1 drives cotranscription of edd and eda and is induced by growth on gluconate (7); P1 appears to be controlled by GntR (20,21,24), although binding of GntR to the P1 regulatory region was not established prior to this work. A second promoter, P2, is located within the terminal one-third of the edd coding sequence, and the closely spaced third and fourth putative promoters (P3 and P4) are located at the distal end of the edd coding region (7). High basal expression is characteristic of eda (20). Although it is known that the P2,...

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Molecular characterization of the Entner-Doudoroff pathway in Escherichia coli: sequence analysis and localization of promoters for the edd-eda operon

Cited by 83 publications

References 39 publications

Two genes for carbohydrate catabolism are divergently transcribed from a region of DNA containing the hexC locus in Pseudomonas aeruginosa PAO1

Two genes for carbohydrate catabolism are divergently transcribed from a region of DNA containing the hexC locus in Pseudomonas aeruginosa PAO1

Identification of the Missing Links in Prokaryotic Pentose Oxidation Pathways

Multiple Regulators Control Expression of the Entner-Doudoroff Aldolase (Eda) of Escherichia coli

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