NH3-dependent NAD+synthetase fromBacillus subtilisat 1 Å resolution

Symerský, J.; Devedjiev, Y.; Moore, Karen; Brouillette, Christie G.; DeLucas, L.J.

doi:10.1107/s0907444902006698

Cited by 32 publications

(24 citation statements)

References 20 publications

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“…The data presented herein for ecoNADS, including the conservation of the mode of substrate and metal ion binding, are in general agreement with the reaction mechanism described for bsuNADS (16), suggesting that it is conserved for bacterial NADSs. However, our ensemble of ecoNADS structures allows us to expand the mechanism to structural changes taking place in the enzymes upon substrate conversion.…”

Section: Resultssupporting

confidence: 85%

“…Subsequently, the picture was expanded by co-crystallizing bsu-NADS with NAAD, AMP ϩ PP i (a combination of NAAD and ATP), and also with the ATP analog AMP-CPP (15). More recently, a 1.0-Å resolution structure of bsuNADS in complex with the NAD-adenylate intermediate has been described (16). The combined structural studies unraveled the overall architecture of a NADS and delineated its mode of substrate binding.…”

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confidence: 99%

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Structures of Escherichia coli NAD Synthetase with Substrates and Products Reveal Mechanistic Rearrangements

Jauch

Humm

Huber

et al. 2005

Journal of Biological Chemistry

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Nicotinamide adenine dinucleotide synthetases (NADS) catalyze the amidation of nicotinic acid adenine dinucleotide (NAAD) to yield the enzyme cofactor nicotinamide adenine dinucleotide (NAD). Here we describe the crystal structures of the ammonia-dependent homodimeric NADS from Escherichia coli alone and in complex with natural substrates and with the reaction product NAD. The structures disclosed two NAAD/NAD binding sites at the dimer interface and an adenosine triphosphate (ATP) binding site within each subunit. Comparison with the Bacillus subtilis NADS showed pronounced chemical differences in the NAAD/NAD binding sites and less prominent differences in the ATP binding pockets. In addition, the E. coli NADS structures revealed unexpected dynamical rearrangements in the NAAD/NAD binding pocket upon NAAD-to-NAD conversion, which define a catalysis state and a substrate/product exchange state. The two states are adopted by concerted movement of the nicotinysyl moieties of NAAD and NAD, Phe-170, and residues 224-228, which may be triggered by differential coordination of a magnesium ion to NAAD and NAD. Phylogenetic structure comparisons suggest that the present results are relevant for designing species-specific antibiotics.

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Section: Resultssupporting

confidence: 85%

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confidence: 99%

Structures of Escherichia coli NAD Synthetase with Substrates and Products Reveal Mechanistic Rearrangements

Jauch

Humm

Huber

et al. 2005

Journal of Biological Chemistry

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“…In the first step of the reaction, two magnesium ions help stabilise the β and γ phosphate groups of ATP, increasing the electrophilicity of the phosphorus atom of the α phosphate ready for attack by deamino-NAD + . 38,39 The acid and base catalysts in the second and third steps of the reaction are believed to be an ammonium ion and a water molecule, respectively. 3.…”

Section: Regeneration Steps and Non-enzyme-catalysed Stepsmentioning

confidence: 99%

The Chemistry of Protein Catalysis

Holliday

Almonacid

Mitchell

et al. 2007

Journal of Molecular Biology

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“…In this study, four commercial compound databases were filtered according to Lipinski's rule of 5 using Tripos' program Unity: Maybridge (58,650 after filtering), ChemBridge (404,132), Tripos' LeadQuest (72,660), and ComGenex (82,737). Because these docking studies predate our solution of the crystal structure of B. anthracis NADs (PDB code 2PZB),18 the highest available resolution crystal structure of B. subtilis NADs,19 reported by our group, was utilized for docking (PDB code 1KQP19). The crystal structures of B. anthracis and B. subtilis NADs reveal that the binding sites are nearly identical, with all active site residues being conserved 18…”

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confidence: 99%

Virtual screening to identify lead inhibitors for bacterial NAD synthetase (NADs)

Moro

Yang

Kane

et al. 2009

Bioorganic & Medicinal Chemistry Letters

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Virtual screening was employed to identify new drug-like inhibitors of NAD synthetase (NADs) as antibacterial agents. Four databases of commercially available compounds were docked against three subsites of the NADs active site using FlexX in conjunction with CScore. Over 200 commercial compounds were purchased and evaluated in enzyme inhibition and antibacterial assays. 18 compounds inhibited NADs at or below 100 μM (7.6% hit rate), and two were selected for future SAR studies.With the increasing threat of pathogens, such as Bacillus anthracis, being used as bioweapons, 1 and the rise in the incidence of multi-drug resistant bacteria, 2 the need for new antibiotics that act at novel targets has never been greater. Previous studies within this group 3-5 have revealed that inhibition of one such target, the amidotransferase enzyme nicotinamide adenine dinucleotide (NAD) synthetase (NADs), could hinder both spore outgrowth and vegetative growth, which would provide antibacterial action at two different steps in the bacterial life cycle. 6-10The first class of NADs inhibitors designed by this group consisted of tethered dimers that contain two hydrophobic groups linked by a polymethylene tether, and a positively charged nitrogen on one end. 3-5 These inhibitors were antibacterial, and there was a correlation between the potencies of enzyme inhibition and antibacterial effects. However, the permanent positive charge and detergent-like properties of this class of compounds were unattractive for further drug development. 11,12 More drug-like lead inhibitors were, therefore, sought.Virtual screening of compound databases using the detailed structure of the drug target can serve to greatly enhance success in the lead discovery process. 13-17 Here we use the in silico screening program FlexX 1.20.1(BiosolveIT GmbH ® ) for the virtual screening of commercially available compounds within the catalytic site of NADs to identify new classes *Corresponding author. Tel. +1 205 934 8288; FAX +1 205 934 2543. Email address: wbrou@uab.edu (W. Brouillette). Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.Supplementary Data Supplementary data, including (a) structures of compounds from Table 1 not shown in Table 2, (b) graphical representations of the binding sites used and poses of selected docked ligands, and (c) a sample HPLC-chromatogram used in the enzyme assay, can be found in the online version. NADs is a large homodimer of approximately 60 kDa that contains two identical binding sites. The crystal structure of the protein from B. subtilis reveals two identical long, linear bind...

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NH₃-dependent NAD⁺synthetase fromBacillus subtilisat 1 Å resolution

Cited by 32 publications

References 20 publications

Structures of Escherichia coli NAD Synthetase with Substrates and Products Reveal Mechanistic Rearrangements

Structures of Escherichia coli NAD Synthetase with Substrates and Products Reveal Mechanistic Rearrangements

The Chemistry of Protein Catalysis

Virtual screening to identify lead inhibitors for bacterial NAD synthetase (NADs)

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