Control of aromatic biosynthesis: The multiplicity of 7-phospho-2-oxo-3-deoxy-d-arabino-heptonate d-erythrose-4-phosphate-lyase (pyruvate-phosphorylating) in Eschetichia coli W

Doy, Colin H.; Brown, Keith

doi:10.1016/0304-4165(65)90343-0

Cited by 110 publications

(30 citation statements)

References 15 publications

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“…In some cases, the cell synthesizes two or more such enzymes simultaneously, as is the case for the three aromatic biosynthesis isoenzymes which catalyze the D-arabino heptulosonate-7-phosphate synthetase reaction (7).…”

Section: Resultsmentioning

confidence: 99%

A novel L-serine deaminase activity in Escherichia coli K-12

Newman

1991

J Bacteriol

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We demonstrate here that Escherichia coli K-12 synthesizes two different L-serine deaminases (L-SD) catalyzing the nonoxidative deamination of L-serine to pyruvate, one coded for by the previously described sdaA gene and a second, hitherto undescribed enzyme which we call L-SD2. A strain carrying a null mutation in sdaA made no detectable L-SD in minimal medium, but had activity in Luria broth. We describe a mutation, sdaX, which affects the regulation of L-SD2 and permits its expression in minimal medium, and an insertion mutation, sdaB, which abolishes L-SD2 activity completely. Both mutations lie near 60.5 min on the E. coli genetic map. The two L-SD enzymes have similar enzyme parameters, and both require posttranslational activation.Escherichia coli K-12, when grown in glucose-minimal medium, makes an enzyme, L-serine deaminase (L-SD), which is further induced in media with glycine and L-leucine added (15,24). This enzyme is even further induced by growth in Luria broth (LB) and is subject to complex regulation by at least two global regulatory systems (17,23). The enzyme is coded by the sdaA gene, which has been cloned and sequenced (28).L-SD activity is also affected by posttranslational control. L-SD is synthesized in an inactive form in E. coli (20). Several mutants have been isolated that are unable to make an active L-SD but make a protein that can be activated in vitro (20).The number of environmental factors and the variety of mechanisms affecting L-SD activity suggest that its metabolic role is important to the cell. Nonetheless, mutants deficient in L-SD show very little difference in phenotype from their parent strain (22, 28).We were surprised to find that, when grown in LB, a strain deficient in L-SD1 still had a considerable amount of L-SD activity, and we proposed that there might be a second L-SD, the synthesis of which was regulated differently from that of the first (22).In this work, we demonstrate the existence of this second enzyme in a strain carrying a null mutation in sdaA. L-SD2 is similar to L-SD1 in many of its parameters, but it clearly is different in structure and is encoded by a separate gene. MATERIALS AND METHODSCultures. The strains used in this study, all derivatives of E. coli K-12, are described in Table 1, as are the bacteriophages and plasmids.Growth conditions. The minimal medium used has been described previously (22). Because strains CU1008 and MEW1 and all their derivatives carry a deletion in ilvA, L-isoleucine and L-valine (50 p.g/ml each) were added to all media. When cells were grown for L-SD assays in LB, 0.5% glucose was added to completely repress the biodegradative L-threonine deaminase.SGL medium. Medium with a combination of L-serine, glycine, and L-leucine as the only carbon sources other than L-isoleucine and L-valine is called SGL medium, and strains * Corresponding author. unable to grow on this medium are termed SGL-(22). L-Serine, glycine, and L-leucine were usually provided at 2,000, 300, and 300 ,ug/ml, respectively.

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Section: Resultsmentioning

confidence: 99%

A novel L-serine deaminase activity in Escherichia coli K-12

Newman

1991

J Bacteriol

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“…This product is then visualized by formation of a chromophore upon the addition of thiobarbituric acid. The method described below is a variation of the method of Weissbach and Hurwitz (28) and modified by Doy and Brown (29).…”

Section: Dahp Synthase Assays (I)mentioning

confidence: 99%

Intracellular Hydrogen Peroxide and Superoxide Poison 3-Deoxy-D-Arabinoheptulosonate 7-Phosphate Synthase, the First Committed Enzyme in the Aromatic Biosynthetic Pathway of Escherichia coli

Sobota

Imlay

2014

Journal of Bacteriology

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In Escherichia coli, aromatic compound biosynthesis is the process that has shown the greatest sensitivity to hydrogen peroxide stress. This pathway has long been recognized to be sensitive to superoxide as well, but the molecular target was unknown. Feeding experiments indicated that the bottleneck lies early in the pathway, and the suppressive effects of fur mutations and manganese supplementation suggested the involvement of a metalloprotein. The 3-deoxy-D-arabinoheptulosonate 7-phosphate synthase (DAHP synthase) activity catalyzes the first step in the pathway, and it is provided by three isozymes known to rely upon a divalent metal. This activity progressively declined when cells were stressed with either oxidant. The purified enzyme was activated more strongly by ferrous iron than by other metals, and only this metalloform could be inactivated by hydrogen peroxide or superoxide. We infer that iron is the prosthetic metal in vivo. Both oxidants displace the iron atom from the enzyme. In peroxide-stressed cells, the enzyme accumulated as an apoprotein, potentially with an oxidized cysteine residue. In superoxidestressed cells, the enzyme acquired a nonactivating zinc ion in its active site, an apparent consequence of the repeated ejection of iron. Manganese supplementation protected the activity in both cases, which matches the ability of manganese to metallate the enzyme and to provide substantial oxidant-resistant activity. DAHP synthase thus belongs to a family of mononuclear iron-containing enzymes that are disabled by oxidative stress. To date, all the intracellular injuries caused by physiological doses of these reactive oxygen species have arisen from the oxidation of reduced iron centers.

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“…In Escherichia coli and other enteric bacteria there are three DAHPS isoenzymes (5). Although catalyzing the same reaction (i.e., the condensation of erythrose-4-phosphate and phosphoenolpyruvate [PEP] to form 3-deoxy-D-arabino-heptulosonate 7-phosphate [DAHP]), each isoenzyme is feedback regulated by a different aromatic amino acid.…”

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confidence: 99%

Mutational analysis of the catalytic and feedback sites of the tryptophan-sensitive 3-deoxy-D-arabino-heptulosonate-7-phosphate synthase of Escherichia coli

1988

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The nucleotide sequence of aroH, the structural gene for the tryptophan-sensitive 3-deoxy-D-arabinoheptulosonate-7-phosphate synthase [DAHPS(Trp)], is presented, and the deduced amino acid sequence of AroH is compared with that of the tyrosine-sensitive (AroF) and phenylalanine-sensitive (AroG) DAHPS isoenzymes, The high degree of sequence similarity among the three isoenzymes strongly indicates that they have a common evolutionary origin. In vitro chemical mutagenesis of the cloned aroH gene was used to identify residues and regions of the polypeptide essential for catalytic activity and for tryptophan feedback regulation. Missense mutations leading either to loss of catalytic activity or to feedback resistance were found interspersed throughout the polypeptide, suggesting overlapping catalytic and regulatory sites in DAHPS(Trp). We conclude that the specfficity of feedback regulation of the isoenzymes was probably acquired by the duplication and divergent evolution of an ancestral gene, rather than by domain recruitment.In bacteria and plants, 3-deoxy-D-arabino-heptulosonate 7-phosphate synthase (DAHPS) (EC 4.1.2.15) catalyzes the first committed step in the pathway that leads to the biosynthesis of aromatic acids and vitamins. In Escherichia coli and other enteric bacteria there are three DAHPS isoenzymes (5). Although catalyzing the same reaction (i.e., the condensation of erythrose-4-phosphate and phosphoenolpyruvate [PEP] to form 3-deoxy-D-arabino-heptulosonate 7-phosphate [DAHP]), each isoenzyme is feedback regulated by a different aromatic amino acid. The structural genes aroF, aroG, and aroH encode the tyrosine (Tyr)-, phenylalanine (Phe)-, and tryptophan (Trp

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Control of aromatic biosynthesis: The multiplicity of 7-phospho-2-oxo-3-deoxy-d-arabino-heptonate d-erythrose-4-phosphate-lyase (pyruvate-phosphorylating) in Eschetichia coli W

Cited by 110 publications

References 15 publications

A novel L-serine deaminase activity in Escherichia coli K-12

A novel L-serine deaminase activity in Escherichia coli K-12

Intracellular Hydrogen Peroxide and Superoxide Poison 3-Deoxy-D-Arabinoheptulosonate 7-Phosphate Synthase, the First Committed Enzyme in the Aromatic Biosynthetic Pathway of Escherichia coli

Mutational analysis of the catalytic and feedback sites of the tryptophan-sensitive 3-deoxy-D-arabino-heptulosonate-7-phosphate synthase of Escherichia coli

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