Dataset Information

Ligand binding induces an ammonia channel in 2-amino-2-desoxyisochorismate (ADIC) synthase PhzE.

ABSTRACT: PhzE utilizes chorismate and glutamine to synthesize 2-amino-2-desoxyisochorismate (ADIC) in the first step of phenazine biosynthesis. The PhzE monomer contains both a chorismate-converting menaquinone, siderophore, tryptophan biosynthesis (MST) and a type 1 glutamine amidotransferase (GATase1) domain connected by a 45-residue linker. We present here the crystal structure of PhzE from Burkholderia lata 383 in a ligand-free open and ligand-bound closed conformation at 2.9 and 2.1 Å resolution, respectively. PhzE arranges in an intertwined dimer such that the GATase1 domain of one chain provides NH(3) to the MST domain of the other. This quaternary structure was confirmed by small angle x-ray scattering. Binding of chorismic acid, which was found converted to benzoate and pyruvate in the MST active centers of the closed form, leads to structural rearrangements that establish an ammonia transport channel approximately 25 Å in length within each of the two MST/GATase1 functional units of the dimer. The assignment of PhzE as an ADIC synthase was confirmed by mass spectrometric analysis of the product, which was also visualized at 1.9 Å resolution by trapping in crystals of an inactive mutant of PhzD, an isochorismatase that catalyzes the subsequent step in phenazine biosynthesis. Unlike in some of the related anthranilate synthases, no allosteric inhibition was observed in PhzE. This can be attributed to a tryptophan residue of the protein blocking the potential regulatory site. Additional electron density in the GATase1 active center was identified as zinc, and it was demonstrated that Zn(2+), Mn(2+), and Ni(2+) reduce the activity of PhzE.

SUBMITTER: Li QA

PROVIDER: S-EPMC3093893 | biostudies-literature | 2011 May

REPOSITORIES: biostudies-literature

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Ligand binding induces an ammonia channel in 2-amino-2-desoxyisochorismate (ADIC) synthase PhzE.

Li Qi-Ang QA Mavrodi Dmitri V DV Thomashow Linda S LS Roessle Manfred M Blankenfeldt Wulf W

The Journal of biological chemistry 20110329 20

PhzE utilizes chorismate and glutamine to synthesize 2-amino-2-desoxyisochorismate (ADIC) in the first step of phenazine biosynthesis. The PhzE monomer contains both a chorismate-converting menaquinone, siderophore, tryptophan biosynthesis (MST) and a type 1 glutamine amidotransferase (GATase1) domain connected by a 45-residue linker. We present here the crystal structure of PhzE from Burkholderia lata 383 in a ligand-free open and ligand-bound closed conformation at 2.9 and 2.1 Å resolution, re ...[more]

PMID: 21454481

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Project description:Aerial ammonia exposure leads to tissue damage and metabolic dysfunction. However, it is unclear how different organs are coordinated to defend against aerial ammonia exposure. Twenty-four pigs were randomly divided into 4 groups, exposed to 0, 10, 25 or 35 mg/m3 ammonia respectively for 25 d. After above 25 mg/m3 ammonia exposure, decreased aspartate (P = 0.016), glutamate (P = 0.030) and increased ornithine (P = 0.002) were found in the ammonia-removing liver, and after high ammonia (35 mg/m3) exposure, glutamine synthetase (GS) expression was increased (P = 0.012). An increased glutamate (P = 0.004) and decreased glutaminase (GLS) expression (P = 0.083) were observed in the lungs after high ammonia exposure. There was also an increasing trend of glutamine in the kidneys after high ammonia exposure (P = 0.066). For branched-chain amino acid (BCAA) catabolism, high ammonia exposure increased BCAA content in both the lungs and muscle (P < 0.05), whereas below 25 mg/m3 ammonia exposure increased BCAA only in the lungs (P < 0.05). The expression of BCAA transaminase (BCAT1/2) and dehydrogenase complex (BCKDHA/B and DBT) were inhibited to a varying degree in the liver, lungs and muscle after above 25 mg/m3 ammonia exposure, especially high ammonia exposure. The expression of BCKDH complex and glutamate-glutamine metabolism-related genes were highly expressed in the liver, followed by the lungs and muscle (P < 0.01), whereas the BCAT2 expression was highest in the lungs (P = 0.002). Altogether, low ammonia exposure sufficed to evoke the urea cycle to detoxify ammonia in the liver. The process of ammonia removal in the liver and potential ability of the lungs to detoxify ammonia were enhanced with increasing ammonia. Furthermore, high ammonia exposure impaired the BCAA catabolism and decreased the transcripts of the BCAA catabolism-related enzymes, resulting in high BCAA content in extrahepatic tissues. Therefore, with aerial ammonia increasing, an increased urea cycle and glutamine synthesis were ammonia defensive strategies, and high ammonia exposure impaired the BCAA catabolism.

Dataset Information

Ligand binding induces an ammonia channel in 2-amino-2-desoxyisochorismate (ADIC) synthase PhzE.

Publications

Ligand binding induces an ammonia channel in 2-amino-2-desoxyisochorismate (ADIC) synthase PhzE.

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