DAHP Synthase

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Paul J Berti - One of the best experts on this subject based on the ideXlab platform.

  • eliminating competition characterizing and eliminating competitive binding at separate sites between DAHP Synthase s essential metal ion and the inhibitor DAHP oxime
    Biochemistry, 2018
    Co-Authors: Maren Heimhalt, Shan Jiang, Paul J Berti
    Abstract:

    3-Deoxy-d-arabinoheptulosonate 7-phosphate (DAHP) oxime is a transition state mimic inhibitor of bacterial DAHP Synthase, with Ki = 1.5 μM and a residence time of tR = 83 min. Unexpectedly, DAHP oxime inhibition is competitive with respect to the essential metal ion, Mn2+, even though the inhibitor and metal ion do not occupy the same physical space in the active site. This is problematic because DAHP Synthase is activated by multiple divalent metal cations, some of which have significant intracellular concentrations and some of which dissociate slowly. The nature of DAHP oxime’s competition with the metal ion was investigated. Inhibition shifted from metal-competitive at physiological pH to metal-noncompetitive at pH > 8.7 in response to deprotonation of the Cys61 side chain. The modes of inhibition of DAHP Synthase mutants and inhibitor fragments demonstrated that metal-competitive inhibition arose from interactions between Mn2+, DAHP oxime’s O4 hydroxyl group, and the Cys61 and Asp326 side chains. The ...

  • Eliminating Competition: Characterizing and Eliminating Competitive Binding at Separate Sites between DAHP Synthase’s Essential Metal Ion and the Inhibitor DAHP Oxime
    2018
    Co-Authors: Maren Heimhalt, Shan Jiang, Paul J Berti
    Abstract:

    3-Deoxy-d-arabinoheptulosonate 7-phosphate (DAHP) oxime is a transition state mimic inhibitor of bacterial DAHP Synthase, with Ki = 1.5 μM and a residence time of tR = 83 min. Unexpectedly, DAHP oxime inhibition is competitive with respect to the essential metal ion, Mn2+, even though the inhibitor and metal ion do not occupy the same physical space in the active site. This is problematic because DAHP Synthase is activated by multiple divalent metal cations, some of which have significant intracellular concentrations and some of which dissociate slowly. The nature of DAHP oxime’s competition with the metal ion was investigated. Inhibition shifted from metal-competitive at physiological pH to metal-noncompetitive at pH > 8.7 in response to deprotonation of the Cys61 side chain. The modes of inhibition of DAHP Synthase mutants and inhibitor fragments demonstrated that metal-competitive inhibition arose from interactions between Mn2+, DAHP oxime’s O4 hydroxyl group, and the Cys61 and Asp326 side chains. The majority of potent DAHP Synthase inhibitors in the literature possess a 4-hydroxyl group. Removing it could avoid metal-competitive inhibition and avoid them being outcompeted by metal ions in vivo

  • linear free energy relationship analysis of transition state mimicry by 3 deoxy d arabino heptulosonate 7 phosphate DAHP oxime a DAHP Synthase inhibitor and phosphate mimic
    Biochemistry, 2017
    Co-Authors: Naresh Balachandran, Paul J Berti
    Abstract:

    3-Deoxy-d-arabino-heptulosonate-7-phosphate (DAHP) Synthase catalyzes an aldol-like reaction of phosphoenolpyruvate (PEP) with erythrose 4-phosphate (E4P) to form DAHP in the first step of the shikimate biosynthetic pathway. DAHP oxime, in which an oxime replaces the ketone, is a potent inhibitor, with Ki = 1.5 μM. Linear free energy relationship (LFER) analysis of DAHP oxime inhibition using DAHP Synthase mutants revealed an excellent correlation between transition state stabilization and inhibition. The equations of LFER analysis were rederived to formalize the possibility of proportional, rather than equal, changes in the free energies of transition state stabilization and inhibitor binding, in accord with the fact that the majority of LFER analyses in the literature demonstrate nonunity slopes. A slope of unity, m = 1, indicates that catalysis and inhibitor binding are equally sensitive to perturbations such as mutations or modified inhibitor/substrate structures. Slopes 1 indicate that inhibit...

  • Linear Free Energy Relationship Analysis of Transition State Mimicry by 3‑Deoxy‑d-arabino-heptulosonate-7-phosphate (DAHP) Oxime, a DAHP Synthase Inhibitor and Phosphate Mimic
    2017
    Co-Authors: Naresh Balachandran, Paul J Berti
    Abstract:

    3-Deoxy-d-arabino-heptulosonate-7-phosphate (DAHP) Synthase catalyzes an aldol-like reaction of phosphoenolpyruvate (PEP) with erythrose 4-phosphate (E4P) to form DAHP in the first step of the shikimate biosynthetic pathway. DAHP oxime, in which an oxime replaces the ketone, is a potent inhibitor, with Ki = 1.5 μM. Linear free energy relationship (LFER) analysis of DAHP oxime inhibition using DAHP Synthase mutants revealed an excellent correlation between transition state stabilization and inhibition. The equations of LFER analysis were rederived to formalize the possibility of proportional, rather than equal, changes in the free energies of transition state stabilization and inhibitor binding, in accord with the fact that the majority of LFER analyses in the literature demonstrate nonunity slopes. A slope of unity, m = 1, indicates that catalysis and inhibitor binding are equally sensitive to perturbations such as mutations or modified inhibitor/substrate structures. Slopes 1 indicate that inhibitor binding is less sensitive or more sensitive, respectively, to perturbations than is catalysis. LFER analysis using the tetramolecular specificity constant, that is, plotting log­(KM,MnKM,PEPKM,E4P/kcat) versus log­(Ki), revealed a slope, m, of 0.34, with r2 = 0.93. This provides evidence that DAHP oxime is mimicking the first irreversible transition state of the DAHP Synthase reaction, presumably phosphate departure from the tetrahedral intermediate. This is evidence that the oxime group can act as a functional, as well as structural, mimic of phosphate groups

  • potent inhibition of 3 deoxy d arabinoheptulosonate 7 phosphate DAHP Synthase by DAHP oxime a phosphate group mimic
    Biochemistry, 2016
    Co-Authors: Naresh Balachandran, Maren Heimhalt, Peter Liuni, Derek J Wilson, Murray S Junop, Paul J Berti
    Abstract:

    3-Deoxy-d-arabinoheptulosonate-7-phosphate (DAHP) Synthase catalyzes the first step in the shikimate pathway. It catalyzes an aldol-like reaction of phosphoenolpyruvate (PEP) with erythrose 4-phosphate (E4P) to form DAHP. The kinetic mechanism was rapid equilibrium sequential ordered ter ter, with the essential divalent metal ion, Mn2+, binding first, followed by PEP and E4P. DAHP oxime, in which an oxime group replaces the keto oxygen, was a potent inhibitor, with Ki = 1.5 ± 0.4 μM, though with residual activity at high inhibitor concentrations. It displayed slow-binding inhibition with a residence time, tR, of 83 min. The crystal structure revealed that the oxime functional group, combined with two crystallographic waters, bound at the same location in the catalytic center as the phosphate group of the tetrahedral intermediate. DAHP Synthase has a dimer-of-dimers homotetrameric structure, and DAHP oxime bound to only one subunit of each tight dimer. Inhibitor binding was competitive with respect to all ...

Naresh Balachandran - One of the best experts on this subject based on the ideXlab platform.

  • linear free energy relationship analysis of transition state mimicry by 3 deoxy d arabino heptulosonate 7 phosphate DAHP oxime a DAHP Synthase inhibitor and phosphate mimic
    Biochemistry, 2017
    Co-Authors: Naresh Balachandran, Paul J Berti
    Abstract:

    3-Deoxy-d-arabino-heptulosonate-7-phosphate (DAHP) Synthase catalyzes an aldol-like reaction of phosphoenolpyruvate (PEP) with erythrose 4-phosphate (E4P) to form DAHP in the first step of the shikimate biosynthetic pathway. DAHP oxime, in which an oxime replaces the ketone, is a potent inhibitor, with Ki = 1.5 μM. Linear free energy relationship (LFER) analysis of DAHP oxime inhibition using DAHP Synthase mutants revealed an excellent correlation between transition state stabilization and inhibition. The equations of LFER analysis were rederived to formalize the possibility of proportional, rather than equal, changes in the free energies of transition state stabilization and inhibitor binding, in accord with the fact that the majority of LFER analyses in the literature demonstrate nonunity slopes. A slope of unity, m = 1, indicates that catalysis and inhibitor binding are equally sensitive to perturbations such as mutations or modified inhibitor/substrate structures. Slopes 1 indicate that inhibit...

  • Linear Free Energy Relationship Analysis of Transition State Mimicry by 3‑Deoxy‑d-arabino-heptulosonate-7-phosphate (DAHP) Oxime, a DAHP Synthase Inhibitor and Phosphate Mimic
    2017
    Co-Authors: Naresh Balachandran, Paul J Berti
    Abstract:

    3-Deoxy-d-arabino-heptulosonate-7-phosphate (DAHP) Synthase catalyzes an aldol-like reaction of phosphoenolpyruvate (PEP) with erythrose 4-phosphate (E4P) to form DAHP in the first step of the shikimate biosynthetic pathway. DAHP oxime, in which an oxime replaces the ketone, is a potent inhibitor, with Ki = 1.5 μM. Linear free energy relationship (LFER) analysis of DAHP oxime inhibition using DAHP Synthase mutants revealed an excellent correlation between transition state stabilization and inhibition. The equations of LFER analysis were rederived to formalize the possibility of proportional, rather than equal, changes in the free energies of transition state stabilization and inhibitor binding, in accord with the fact that the majority of LFER analyses in the literature demonstrate nonunity slopes. A slope of unity, m = 1, indicates that catalysis and inhibitor binding are equally sensitive to perturbations such as mutations or modified inhibitor/substrate structures. Slopes 1 indicate that inhibitor binding is less sensitive or more sensitive, respectively, to perturbations than is catalysis. LFER analysis using the tetramolecular specificity constant, that is, plotting log­(KM,MnKM,PEPKM,E4P/kcat) versus log­(Ki), revealed a slope, m, of 0.34, with r2 = 0.93. This provides evidence that DAHP oxime is mimicking the first irreversible transition state of the DAHP Synthase reaction, presumably phosphate departure from the tetrahedral intermediate. This is evidence that the oxime group can act as a functional, as well as structural, mimic of phosphate groups

  • potent inhibition of 3 deoxy d arabinoheptulosonate 7 phosphate DAHP Synthase by DAHP oxime a phosphate group mimic
    Biochemistry, 2016
    Co-Authors: Naresh Balachandran, Maren Heimhalt, Peter Liuni, Derek J Wilson, Murray S Junop, Paul J Berti
    Abstract:

    3-Deoxy-d-arabinoheptulosonate-7-phosphate (DAHP) Synthase catalyzes the first step in the shikimate pathway. It catalyzes an aldol-like reaction of phosphoenolpyruvate (PEP) with erythrose 4-phosphate (E4P) to form DAHP. The kinetic mechanism was rapid equilibrium sequential ordered ter ter, with the essential divalent metal ion, Mn2+, binding first, followed by PEP and E4P. DAHP oxime, in which an oxime group replaces the keto oxygen, was a potent inhibitor, with Ki = 1.5 ± 0.4 μM, though with residual activity at high inhibitor concentrations. It displayed slow-binding inhibition with a residence time, tR, of 83 min. The crystal structure revealed that the oxime functional group, combined with two crystallographic waters, bound at the same location in the catalytic center as the phosphate group of the tetrahedral intermediate. DAHP Synthase has a dimer-of-dimers homotetrameric structure, and DAHP oxime bound to only one subunit of each tight dimer. Inhibitor binding was competitive with respect to all ...

  • TITLE: DAHP Oxime: A Transition State Mimic Inhibitor Of
    2016
    Co-Authors: Naresh Balachandran
    Abstract:

    The rise of bacterial infections and increase of antibiotic resistant bacteria has become a major problem in the treatment of bacterial infections. The use and overuse of antibiotics, and the inherent ability of bacteria to adapt to their environment, have lead to the emergence of strains that are resistant to all antibiotics. Ideally, new targets for antibacterial drug therapy would be essential to the virulence of most or all bacteria. That is, antibiotics exploiting these targets would have broad spectrum activity. 3-Deoxy-D-arabinoheptulosonate-7-phosphate (DAHP) Synthase could be such a target. This enzyme catalyzes the condensation of erythrose 4-phosphate (E4P) and phoshoenolpyruvate (PEP) to form DAHP. The DAHP Synthase-catalyzed reaction is the first committed step in the shikimic acid biosynthetic pathway leading to the aromatic amino acids and other secondary metabolites in all bacteria and some parasites. Inhibition of this enzyme would lead to a depletion of aromatic amino acids within the cell, halting new protein synthesis and killing the cells. Our lab has developed a transitio

  • Potent Inhibition of 3‑Deoxy‑d‑arabinoheptulosonate-7-phosphate (DAHP) Synthase by DAHP Oxime, a Phosphate Group Mimic
    2016
    Co-Authors: Naresh Balachandran, Maren Heimhalt, Peter Liuni, Derek J Wilson, Murray S Junop, Paul J Berti
    Abstract:

    3-Deoxy-d-arabinoheptulosonate-7-phosphate (DAHP) Synthase catalyzes the first step in the shikimate pathway. It catalyzes an aldol-like reaction of phosphoenolpyruvate (PEP) with erythrose 4-phosphate (E4P) to form DAHP. The kinetic mechanism was rapid equilibrium sequential ordered ter ter, with the essential divalent metal ion, Mn2+, binding first, followed by PEP and E4P. DAHP oxime, in which an oxime group replaces the keto oxygen, was a potent inhibitor, with Ki = 1.5 ± 0.4 μM, though with residual activity at high inhibitor concentrations. It displayed slow-binding inhibition with a residence time, tR, of 83 min. The crystal structure revealed that the oxime functional group, combined with two crystallographic waters, bound at the same location in the catalytic center as the phosphate group of the tetrahedral intermediate. DAHP Synthase has a dimer-of-dimers homotetrameric structure, and DAHP oxime bound to only one subunit of each tight dimer. Inhibitor binding was competitive with respect to all three substrates in the subunits to which it bound. DAHP oxime did not overlap with the metal binding site, so the cause of their mutually exclusive binding was not clear. Similarly, there was no obvious structural reason for inhibitor binding in only two subunits; however, changes in global hydrogen/deuterium exchange showed large scale changes in protein dynamics upon inhibitor binding. The kcat value for the residual activity at high inhibitor concentrations was 3-fold lower, and the apparent KM,E4P value decreased at least 10-fold. This positive cooperativity of binding between DAHP oxime in subunits B and C, and E4P in subunits A and D appears to be the dominant cause for incomplete inhibition at high inhibitor concentrations. In spite of its lack of obvious structural similarity to phosphate, the oxime and crystallographic waters acted as a small, neutral phosphate mimic

Masaaki Sakuta - One of the best experts on this subject based on the ideXlab platform.

  • effects of conditioned medium on activities of pal chs DAHP Synthase ds co and ds mn and anthocyanin production in suspension cultures of fragaria ananassa
    Plant Science, 2001
    Co-Authors: Tsukasa Mori, Miei Sakurai, Masaaki Sakuta
    Abstract:

    A conditioned medium (CM) prepared from cell suspension cultures of strawberry stimulated anthocyanin synthesis. The effect was significantly (P<0.05) greater than that of synthetic medium (SM), with macronutrient concentrations, carbohydrate concentrations and pH adjusted to those of CM. The activity of 3-deoxy-D-arabino-heptulosonate 7-phosphate (DAHP) Synthase (EC 4.1.2.15) (DS-Mn, DS-Co), phenylalanine ammonia-lyase (PAL, EC 4.3.1.5) and chalcone Synthase (CHS, EC 2.3.1.74) were monitored in the CM- and SM-cultured cells. PAL and CHS activities were found to increase significantly (P<0.05) in the CM-cultured cells. CHS transcript levels were higher in the CM-cultured cells compared to transcript abundance in SM-cultured cells. There was no significant difference in the DS-Mn and DS-Co activities of cells grown in conditioned or synthetic media.

  • changes in pal chs DAHP Synthase ds co and ds mn activity during anthocyanin synthesis in suspension culture of fragaria ananassa
    Plant Cell Tissue and Organ Culture, 2000
    Co-Authors: Tsukasa Mori, Miei Sakurai, Masaaki Sakuta
    Abstract:

    The activity of 3-deoxy-D-arabino-heptulosonate 7-phosphate (DAHP) Synthase (DS-Mn, DS-Co), phenylalanine ammonia-lyase (PAL), and chalcone Synthase (CHS) was monitored at various light intensities (dark, 8.88 μmol m−2 s−1, 88.8 μmol m−2 s−1) using a strawberry cell suspension culture. DS-Mn, PAL, and CHS were found to increase significantly (p>0.05) under light intensitie of 88.8 μmol m−2 s−1 compared to those of 8.88 μmol m−2 s−1 and dark. The activity of DS-Mn, PAL, and CHS were maximum at 88.8 μmol m−2 s−1. Anthocyanin content reached a maximum after 48–60 h of culturing at 88.8 μmol m−2 s−1. DS-Co showed greater activity than DS-Mn during cell culturing, but showed no correlation with anthocyanin production and light intensity. The CHS gene expression was continuous at a light intensity of 88.8 μmol m−2 s−1.

  • changes in the activity of 3 deoxy d arabino heptulosonate 7 phosphate DAHP Synthase in suspension cultured cells of vitis
    Physiologia Plantarum, 1995
    Co-Authors: Nao Suzuki, Masaaki Sakuta, Seki Shimizu, Atsushi Komamine
    Abstract:

    3-Deoxy-D-arabino-heptulosonate 7-phosphate (DAHP) Synthase (EC 4.1.2.15) is the first enzyme in the shikimate pathway, which leads to the biosynthesis of the aromatic amino acids. These amino acids are utilized as precursors for the synthesis of some secondary metabolites. The relationship between the accumulation of anthocyanin and the activity of DAHP Synthase in suspension cultures of Vitis hybrid (Bailey Alicante A) was investigated. The activity of the plastidic isozyme, designated DS-Mn, was very low throughout the culture of cells. However, the activity of the cytosolic isozyme, designated DS-Co, increased transiently and then decreased after transfer of cells to fresh medium, reaching minimum levels during the logarithmic phase. Thereafter, the activity of DS-Co increased rapidly prior to the accumulation of anthocyanin. When phosphate was removed from the culture medium, growth of cells was limited and rapid accumulation of anthocyanin occurred, coincident with the termination of cell division. The activity of phenylalanine ammonia-lyase continued to increase from day 1 and the activity of DS-Co in phosphate-free culture also was 1.6-fold greater than that in the control culture on day 1, while the activity of DS-Mn was unaffected by this treatment. These results suggest a close correlation between the activity of DS-Co and the biosynthesis of anthocyanin.

Ute Krengel - One of the best experts on this subject based on the ideXlab platform.

  • inter enzyme allosteric regulation of chorismate mutase in corynebacterium glutamicum structural basis of feedback activation by trp
    Biochemistry, 2017
    Co-Authors: Daniel Burschowsky, Peter Kast, Helen Vikdal Thorbjornsrud, Joel B Heim, Jūratė Fahrigkamarauskaitė, Kathrin Wurthroderer, Ute Krengel
    Abstract:

    Corynebacterium glutamicum is widely used for the industrial production of amino acids, nucleotides, and vitamins. The shikimate pathway enzymes DAHP Synthase (CgDS, Cg2391) and chorismate mutase (CgCM, Cgl0853) play a key role in the biosynthesis of aromatic compounds. Here we show that CgCM requires the formation of a complex with CgDS to achieve full activity, and that both CgCM and CgDS are feedback regulated by aromatic amino acids binding to CgDS. Kinetic analysis showed that Phe and Tyr inhibit CgCM activity by inter-enzyme allostery, whereas binding of Trp to CgDS strongly activates CgCM. Mechanistic insights were gained from crystal structures of the CgCM homodimer, tetrameric CgDS, and the heterooctameric CgCM–CgDS complex, refined to 1.1, 2.5, and 2.2 A resolution, respectively. Structural details from the allosteric binding sites reveal that DAHP Synthase is recruited as the dominant regulatory platform to control the shikimate pathway, similar to the corresponding enzyme complex from Mycobact...

  • Inter-Enzyme Allosteric Regulation of Chorismate Mutase in Corynebacterium glutamicum: Structural Basis of Feedback Activation by Trp
    2017
    Co-Authors: Daniel Burschowsky, Peter Kast, Helen Vikdal Thorbjornsrud, Joel B Heim, Ju̅ratė Fahrig-kamarauskaitė, Kathrin Würth-roderer, Ute Krengel
    Abstract:

    Corynebacterium glutamicum is widely used for the industrial production of amino acids, nucleotides, and vitamins. The shikimate pathway enzymes DAHP Synthase (CgDS, Cg2391) and chorismate mutase (CgCM, Cgl0853) play a key role in the biosynthesis of aromatic compounds. Here we show that CgCM requires the formation of a complex with CgDS to achieve full activity, and that both CgCM and CgDS are feedback regulated by aromatic amino acids binding to CgDS. Kinetic analysis showed that Phe and Tyr inhibit CgCM activity by inter-enzyme allostery, whereas binding of Trp to CgDS strongly activates CgCM. Mechanistic insights were gained from crystal structures of the CgCM homodimer, tetrameric CgDS, and the heterooctameric CgCM–CgDS complex, refined to 1.1, 2.5, and 2.2 Å resolution, respectively. Structural details from the allosteric binding sites reveal that DAHP Synthase is recruited as the dominant regulatory platform to control the shikimate pathway, similar to the corresponding enzyme complex from Mycobacterium tuberculosis

  • remote control by inter enzyme allostery a novel paradigm for regulation of the shikimate pathway
    Journal of Molecular Biology, 2016
    Co-Authors: Steffi Munack, Peter Kast, Severin Sasso, Mats Ökvist, Kathrin Roderer, Jurate Kamarauskaite, Andre Van Eerde, Ute Krengel
    Abstract:

    DAHP Synthase and chorismate mutase catalyze key steps in the shikimate biosynthetic pathway en route to aromatic amino acids. In Mycobacterium tuberculosis, chorismate mutase (MtCM; Rv0948c), located at the branch point toward phenylalanine and tyrosine, has poor activity on its own. However, it is efficiently activated by the first enzyme of the pathway, DAHP Synthase (MtDS; Rv2178c), through formation of a non-covalent MtCM-MtDS complex. Here, we show how MtDS serves as an allosteric platform for feedback regulation of both enzymes, using X-ray crystallography, small-angle X-ray scattering, size-exclusion chromatography, and multi-angle light scattering. Crystal structures of the fully inhibited MtDS and the allosterically down-regulated MtCM-MtDS complex, solved at 2.8 and 2.7A, respectively, reveal how effector binding at the internal MtDS subunit interfaces regulates the activity of MtDS and MtCM. While binding of all three metabolic end products to MtDS shuts down the entire pathway, the binding of phenylalanine jointly with tyrosine releases MtCM from the MtCM-MtDS complex, hence suppressing MtCM activation by 'inter-enzyme allostery'. This elegant regulatory principle, invoking a transient allosteric enzyme interaction, seems to be driven by dynamics and is likely a general strategy used by nature.

  • a novel noncovalent complex of chorismate mutase and DAHP Synthase from mycobacterium tuberculosis protein purification crystallization and x ray diffraction analysis
    Acta Crystallographica Section F-structural Biology and Crystallization Communications, 2009
    Co-Authors: Mats Ökvist, Peter Kast, Severin Sasso, Kathrin Roderer, Ute Krengel
    Abstract:

    Chorismate mutase catalyzes a key step in the shikimate-biosynthetic pathway and hence is an essential enzyme in bacteria, plants and fungi. Mycobacterium tuberculosis contains two chorismate mutases, a secreted and an intracellular one, the latter of which (MtCM; Rv0948c; 90 amino-acid residues; 10 kDa) is the subject of this work. Here are reported the gene expression, purification and crystallization of MtCM alone and of its complex with another shikimate-pathway enzyme, DAHP Synthase (MtDS; Rv2178c; 472 amino-acid residues; 52 kDa), which has been shown to enhance the catalytic efficiency of MtCM. The MtCM–MtDS complex represents the first noncovalent enzyme complex from the common shikimate pathway to be structurally characterized. Soaking experiments with a transition-state analogue are also reported. The crystals of MtCM and the MtCM–MtDS complex diffracted to 1.6 and 2.1 A resolution, respectively.

  • Structure and Function of a Complex between Chorismate Mutase and DAHP Synthase: Efficiency Boost for the Junior Partner.
    The EMBO journal, 2009
    Co-Authors: Severin Sasso, Mats Ökvist, Kathrin Roderer, Marianne Gamper, Giosiana Codoni, Ute Krengel, Peter Kast
    Abstract:

    Chorismate mutase catalyzes a key step in the shikimate biosynthetic pathway towards phenylalanine and tyrosine. Curiously, the intracellular chorismate mutase of Mycobacterium tuberculosis (MtCM; Rv0948c) has poor activity and lacks prominent active-site residues. However, its catalytic efficiency increases >100-fold on addition of DAHP Synthase (MtDS; Rv2178c), another shikimate-pathway enzyme. The 2.35 A crystal structure of the MtCM–MtDS complex bound to a transition-state analogue shows a central core formed by four MtDS subunits sandwiched between two MtCM dimers. Structural comparisons imply catalytic activation to be a consequence of the repositioning of MtCM active-site residues on binding to MtDS. The mutagenesis of the C-terminal extrusion of MtCM establishes conserved residues as part of the activation machinery. The chorismate-mutase activity of the complex, but not of MtCM alone, is inhibited synergistically by phenylalanine and tyrosine. The complex formation thus endows the shikimate pathway of M. tuberculosis with an important regulatory feature. Experimental evidence suggests that such non-covalent enzyme complexes comprising an AroQδ subclass chorismate mutase like MtCM are abundant in the bacterial order Actinomycetales.

John M. Henstrand - One of the best experts on this subject based on the ideXlab platform.