Inhibitors of phosphopantetheine adenylyltransferase

Zhao, Lihua; Allanson, Nigel M.; Thomson, Samantha P.; Maclean, John; Barker, John J.; Primrose, William U.; Tyler, Paul D.; Lewendon, Ann

doi:10.1016/s0223-5234(03)00047-3

Cited by 42 publications

(34 citation statements)

References 4 publications

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“…These include E. coli in complex with several ligands (9,10,25), Thermus thermophilus in complex with PhP (21), Bacillus subtilis in complex with ADP (2), and Mycobacterium tuberculosis (apo-protein) (15). Recently, crystals of Enterococcus faecalis (apo-protein) (13) and Helicobacter pylori (5) have been reported.…”

Section: (Cmentioning

confidence: 99%

Phosphopantetheine Adenylyltransferase from Escherichia coli : Investigation of the Kinetic Mechanism and Role in Regulation of Coenzyme A Biosynthesis

Miller¹,

Ohren²,

Sarver³

et al. 2007

J Bacteriol

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Phosphopantetheine adenylyltransferase (PPAT) from Escherichia coli is an essential hexameric enzyme that catalyzes the penultimate step in coenzyme A (CoA) biosynthesis and is a target for antibacterial drug discovery. The enzyme utilizes Mg-ATP and phosphopantetheine (PhP) to generate dephospho-CoA (dPCoA) and pyrophosphate. When overexpressed in E. coli, PPAT copurifies with tightly bound CoA, suggesting a feedback inhibitory role for this cofactor. Using an enzyme-coupled assay for the forward-direction reaction (dPCoA-generating) and isothermal titration calorimetry, we investigated the steady-state kinetics and ligand binding properties of PPAT. All substrates and products bind the free enzyme, and product inhibition studies are consistent with a random bi-bi kinetic mechanism. CoA inhibits PPAT and is competitive with ATP, PhP, and dPCoA. Previously published structures of PPAT crystallized at pH 5.0 show half-the-sites reactivity for PhP and dPCoA and full occupancy by ATP and CoA. Ligand-binding studies at pH 8.0 show that ATP, PhP, dPCoA, and CoA occupy all six monomers of the PPAT hexamer, although CoA exhibits two thermodynamically distinct binding modes. These results suggest that the half-the-sites reactivity observed in PPAT crystal structures may be pH dependent. In light of previous studies on the regulation of CoA biosynthesis, the PPAT kinetic and ligand binding data suggest that intracellular PhP concentrations modulate the distribution of PPAT monomers between high-and low-affinity CoA binding modes. This model is consistent with PPAT serving as a "backup" regulator of pathway flux relative to pantothenate kinase.

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Section: (Cmentioning

confidence: 99%

Phosphopantetheine Adenylyltransferase from Escherichia coli : Investigation of the Kinetic Mechanism and Role in Regulation of Coenzyme A Biosynthesis

Miller¹,

Ohren²,

Sarver³

et al. 2007

J Bacteriol

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“…Dephospho-coenzyme A synthesis by PPAT is believed to be the rate-limiting step in CoA biosynthesis. Because of the importance of coenzyme A in the citric acid cycle as well as fatty-acid synthesis, coenzyme A biosynthetic proteins are believed to be potential drug targets for infectious disease organisms and PPAT inhibitors have been developed (Zhao et al, 2003).…”

Section: Introductionmentioning

confidence: 99%

Structures of phosphopantetheine adenylyltransferase fromBurkholderia pseudomallei

et al. 2011

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PDB References: PPAT, 3pxu; 3k9w.Phosphopantetheine adenylyltransferase (PPAT) catalyzes the fourth of five steps in the coenzyme A biosynthetic pathway, reversibly transferring an adenylyl group from ATP onto 4 0 -phosphopantetheine to yield dephosphocoenzyme A and pyrophosphate. Burkholderia pseudomallei is a soil-and waterborne pathogenic bacterium and the etiologic agent of melioidosis, a potentially fatal systemic disease present in southeast Asia. Two crystal structures are presented of the PPAT from B. pseudomallei with the expectation that, because of the importance of the enzyme in coenzyme A biosynthesis, they will aid in the search for defenses against this pathogen. A crystal grown in ammonium sulfate yielded a 2.1 Å resolution structure that contained dephospho-coenzyme A with partial occupancy. The overall structure and ligand-binding interactions are quite similar to other bacterial PPAT crystal structures. A crystal grown at low pH in the presence of coenzyme A yielded a 1.6 Å resolution structure in the same crystal form. However, the experimental electron density was not reflective of fully ordered coenzyme A, but rather was only reflective of an ordered 4 0 -diphosphopantetheine moiety.

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“…This analysis is based on the essential requirement of these enzymes for survival and/or virulence and on the lack of homology between bacterial PanK-I enzymes and their mammalian PanK-II counterparts. Development of inhibitors targeting these enzymes is being actively pursued (15,20,49,54,59,60). Among these, the N-substituted alkylpantothenamides have shown the greatest promise as growth inhibitors of both E. coli and S. aureus (15,31,49,59).…”

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confidence: 99%

Crystal Structure of a Type III Pantothenate Kinase: Insight into the Mechanism of an Essential Coenzyme A Biosynthetic Enzyme Universally Distributed in Bacteria

et al. 2006

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Pantothenate kinase (PanK) catalyzes the first step in the five-step universal pathway of coenzyme A (CoA) biosynthesis, a key transformation that generally also regulates the intracellular concentration of CoA through feedback inhibition. A novel PanK protein encoded by the gene coaX was recently identified that is distinct from the previously characterized type I PanK (exemplified by the Escherichia coli coaA-encoded PanK protein) and type II eukaryotic PanKs and is not inhibited by CoA or its thioesters. This type III PanK, or PanK-III, is widely distributed in the bacterial kingdom and accounts for the only known PanK in many pathogenic species, such as Helicobacter pylori, Bordetella pertussis, and Pseudomonas aeruginosa. Here we report the first crystal structure of a type III PanK, the enzyme from Thermotoga maritima (PanK Tm ), solved at 2.0-Å resolution. The structure of PanK Tm reveals that type III PanKs belong to the acetate and sugar kinase/heat shock protein 70/actin (ASKHA) protein superfamily and that they retain the highly conserved active site motifs common to all members of this superfamily. Comparative structural analysis of the PanK Tm active site configuration and mutagenesis of three highly conserved active site aspartates identify these residues as critical for PanK-III catalysis. Furthermore, the analysis also provides an explanation for the lack of CoA feedback inhibition by the enzyme. Since PanK-III adopts a different structural fold from that of the E. coli PanK-which is a member of the "P-loop kinase"superfamily-this finding represents yet another example of convergent evolution of the same biological function from a different protein ancestor.Pantothenate kinase (PanK; EC 2.7.1.33) catalyzes the ATPdependent phosphorylation of pantothenate (vitamin B 5 ) to give 4Ј-phosphopantothenate. This reaction represents the first and committed step in the universal biosynthetic pathway of coenzyme A (CoA) (6, 27, 32). Because CoA is a ubiquitous and essential cofactor in all organisms, genes coding for the five enzymes that make up this pathway-including PanK-are essential for their survival and growth (6,27).Three distinct types of PanK, as differentiated by primary sequence analysis and kinetic properties, have been characterized so far. Type I PanKs (PanK-I) are found exclusively in eubacterial species and are exemplified by the Escherichia coli enzyme encoded by the coaA gene (46, 47). The second type of PanK (PanK-II) is found mainly in eukaryotes, including yeast and various fungi, plants, and mammals (12, 41-43). Interestingly, PanKs from a few gram-positive bacteria, such as Staphylococcus aureus (PanK Sa ) and several bacilli, are also included in this group based on sequence homology, although the bacterial enzymes exhibit certain catalytic characteristics different from their eukaryotic counterparts (15,31). Recently, a third type of PanK (PanK-III) was identified which represents the only known pantothenate kinase activity in many pathogenic bacteria, including Helicobacter pylor...

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Inhibitors of phosphopantetheine adenylyltransferase

Cited by 42 publications

References 4 publications

Phosphopantetheine Adenylyltransferase from Escherichia coli : Investigation of the Kinetic Mechanism and Role in Regulation of Coenzyme A Biosynthesis

Phosphopantetheine Adenylyltransferase from Escherichia coli : Investigation of the Kinetic Mechanism and Role in Regulation of Coenzyme A Biosynthesis

Structures of phosphopantetheine adenylyltransferase fromBurkholderia pseudomallei

Crystal Structure of a Type III Pantothenate Kinase: Insight into the Mechanism of an Essential Coenzyme A Biosynthetic Enzyme Universally Distributed in Bacteria

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