scholarly journals Selective binding of small molecules to Vibrio cholerae DsbA offers a starting point for the design of novel antibacterials

2021 ◽  
Author(s):  
Geqing Wang ◽  
Biswaranjan Mohanty ◽  
Martin Williams ◽  
Bradley Doak ◽  
Rabeb Dhouib ◽  
...  
Keyword(s):  
Vibrio Cholerae ◽  
Crystal Structures ◽  
Small Molecule ◽  
Sodium Taurocholate ◽  
Structural Basis ◽  
Human Pathogens ◽  
X Ray ◽  
X Ray Crystallography ◽  
Hydrophobic Groove ◽  
Starting Point

DsbA enzymes catalyze oxidative folding of proteins that are secreted into the periplasm of Gram-negative bacteria, and they are indispensable for the virulence of human pathogens such as Vibrio cholerae and Escherichia coli. Therefore, targeting DsbA represents an attractive approach to control bacterial virulence. X-ray crystal structures reveal that DsbA enzymes share a similar fold, however, the hydrophobic groove adjacent to the active site, which is implicated in substrate binding, is shorter and flatter in the structure of V. cholerae DsbA (VcDsbA) compared to E. coli DsbA (EcDsbA). The flat and largely featureless nature of this hydrophobic groove is challenging for the development of small molecule inhibitors. Using fragment-based screening approaches, we have identified a novel small molecule, based on the benzimidazole scaffold, that binds to the hydrophobic groove of oxidized VcDsbA with a KD of 446 ± 10 µM. The same benzimidazole compound has ~8-fold selectivity for VcDsbA over EcDsbA and binds to oxidized EcDsbA, with KD > 3.5 mM. We generated a model of the benzimidazole complex with VcDsbA using NMR data but were unable to determine the structure of the benzimidazole bound EcDsbA using either NMR or X-ray crystallography. Therefore, a structural basis for the observed selectivity is unclear. To better understand ligand binding to these two enzymes we crystallized each of them in complex with a known ligand, the bile salt sodium taurocholate. The crystal structures show that taurocholate adopts different binding poses in complex with VcDsbA and EcDsbA, and reveals the protein-ligand interactions that stabilize the different modes of binding. This work highlights the capacity of fragment-based drug discovery to identify inhibitors of challenging protein targets. In addition, it provides a starting point for development of more potent and specific VcDsbA inhibitors that act through a novel anti-virulence mechanism.

scholarly journals ClpP protease activation results from the reorganization of the electrostatic interaction networks at the entrance pores

2019 ◽  
Vol 2 (1) ◽  
Author(s):  
Mark F. Mabanglo ◽  
Elisa Leung ◽  
Siavash Vahidi ◽  
Thiago V. Seraphim ◽  
Bryan T. Eger ◽  
...  
Keyword(s):  
Small Molecule ◽  
Conformational Changes ◽  
Electrostatic Interaction ◽  
Structural Changes ◽  
Structural Heterogeneity ◽  
Interaction Networks ◽  
Structural Basis ◽  
X Ray ◽  
X Ray Crystallography ◽  
Bacterial Cell Death

Abstract Bacterial ClpP is a highly conserved, cylindrical, self-compartmentalizing serine protease required for maintaining cellular proteostasis. Small molecule acyldepsipeptides (ADEPs) and activators of self-compartmentalized proteases 1 (ACP1s) cause dysregulation and activation of ClpP, leading to bacterial cell death, highlighting their potential use as novel antibiotics. Structural changes in Neisseria meningitidis and Escherichia coli ClpP upon binding to novel ACP1 and ADEP analogs were probed by X-ray crystallography, methyl-TROSY NMR, and small angle X-ray scattering. ACP1 and ADEP induce distinct conformational changes in the ClpP structure. However, reorganization of electrostatic interaction networks at the ClpP entrance pores is necessary and sufficient for activation. Further activation is achieved by formation of ordered N-terminal axial loops and reduction in the structural heterogeneity of the ClpP cylinder. Activating mutations recapitulate the structural effects of small molecule activator binding. Our data, together with previous findings, provide a structural basis for a unified mechanism of compound-based ClpP activation.

scholarly journals Insights into the Structures of DNA Damaged by Hydroxyl Radical: Crystal Structures of DNA Duplexes Containing 5-Formyluracil

2010 ◽  
Vol 2010 ◽  
pp. 1-10 ◽  
Author(s):  
Masaru Tsunoda ◽  
Takeshi Sakaue ◽  
Satoko Naito ◽  
Tomoko Sunami ◽  
Naoko Abe ◽  
...  
Keyword(s):  
Crystal Structures ◽  
Dna Polymerase ◽  
Hydroxyl Radicals ◽  
Base Pair ◽  
Structural Basis ◽  
Dna Duplexes ◽  
X Ray ◽  
X Ray Crystallography ◽  
Guanine Base ◽  
New Type

Hydroxyl radicals are potent mutagens that attack DNA to form various base and ribose derivatives. One of the major damaged thymine derivatives is 5-formyluracil (fU), which induces pyrimidine transition during replication. In order to establish the structural basis for such mutagenesis, the crystal structures of two kinds of DNA d(CGCGRATfUCGCG) with R = A/G have been determined by X-ray crystallography. The fU residues form a Watson-Crick-type pair with A and two types of pairs (wobble and reversed wobble) with G, the latter being a new type of base pair between ionized thymine base and guanine base.In silicostructural modeling suggests that the DNA polymerase can accept the reversed wobble pair with G, as well as the Watson-Crick pair with A.

scholarly journals A mixed chirality α-helix in a stapled bicyclic and a linear antimicrobial peptide revealed by X-ray crystallography

2021 ◽  
Author(s):  
Stéphane Baeriswyl ◽  
Hippolyte Personne ◽  
Ivan Di Bonaventura ◽  
Thilo Köhler ◽  
Christian van Delden ◽  
...  
Keyword(s):  
Antimicrobial Peptides ◽  
Antimicrobial Peptide ◽  
Crystal Structures ◽  
X Ray ◽  
X Ray Crystallography ◽  
Α Helix

We report the first X-ray crystal structures of mixed chirality α-helices comprising only natural residues as the example of bicyclic and linear membrane disruptive amphiphilic antimicrobial peptides containing seven l- and four d-residues.

Bis(1-phenyl-1-propyne) complexes of tungsten(II): crystal structures of [WI2(CO)(NCR)(η2-MeC2Ph)2] (R = Me, Et, or Ph)

2001 ◽  
Vol 79 (3) ◽  
pp. 263-271
Author(s):  
Paul K Baker ◽  
Michael GB Drew ◽  
Deborah S Evans
Keyword(s):  
Carbon Monoxide ◽  
Solid State ◽  
Crystal Structures ◽  
Octahedral Geometry ◽  
X Ray ◽  
X Ray Crystallography ◽  
Alkyne Complexes ◽  
First Time

Reaction of [WI2(CO)3(NCMe)2] with two equivalents of 1-phenyl-1-propyne (MeC2Ph) in CH2Cl2, and in the absence of light, gave the bis(1-phenyl-1-propyne) complex [WI2(CO)(NCMe)(η2-MeC2Ph)2] (1) in 77% yield. Treatment of equimolar quantities of 1 and NCR (R = Et, i-Pr, t-Bu, Ph) in CH2Cl2 afforded the nitrile-exchanged products, [WI2(CO)(NCR)(η2-MeC2Ph)2] (2-5) (R = Et (2), i-Pr (3), t-Bu (4), Ph (5)). Complexes 1, 2, and 5 were structurally characterized by X-ray crystallography. All three structures have the same pseudo-octahedral geometry, with the equatorial sites being occupied by cis and parallel alkyne groups, which are trans to the cis-iodo groups. The trans carbon monoxide and acetonitrile ligands occupy the axial sites. In structures 1 and 2, the methyl and phenyl substituents of the 1-phenyl-1-propyne ligands are cis to each other, whereas for the bulkier NCPh complex (5), the methyl and phenyl groups are trans to one another. This is the first time that this arrangement has been observed in the solid state in bis(alkyne) complexes of this type.Key words: bis(1-phenyl-1-propyne), carbonyl, nitrile, diiodo, tungsten(II), crystal structures.

scholarly journals Discovery of Novel Druggable Sites on Zika Virus NS3 Helicase Using X-ray Crystallography-Based Fragment Screening

2018 ◽  
Vol 19 (11) ◽  
pp. 3664 ◽  
Author(s):  
Ali Munawar ◽  
Steven Beelen ◽  
Ahmad Munawar ◽  
Eveline Lescrinier ◽  
Sergei Strelkov
Keyword(s):  
Zika Virus ◽  
Dissociation Constants ◽  
Global Public Health ◽  
Human Pathogens ◽  
Allosteric Site ◽  
Replication Complex ◽  
X Ray ◽  
X Ray Crystallography ◽  
Fragment Screening ◽  
Atomic Precision

The flavivirus family contains several important human pathogens, such as Zika virus (ZIKV), dengue, West Nile, and Yellow Fever viruses, that collectively lead to a large, global disease burden. Currently, there are no approved medicines that can target these viruses. The sudden outbreak of ZIKV infections in 2015–2016 posed a serious threat to global public health. While the epidemic has receded, persistent reservoirs of ZIKV infection can cause reemergence. Here, we have used X-ray crystallography-based screening to discover two novel sites on ZIKV NS3 helicase that can bind drug-like fragments. Both sites are structurally conserved in other flaviviruses, and mechanistically significant. The binding poses of four fragments, two for each of the binding sites, were characterized at atomic precision. Site A is a surface pocket on the NS3 helicase that is vital to its interaction with NS5 polymerase and formation of the flaviviral replication complex. Site B corresponds to a flexible, yet highly conserved, allosteric site at the intersection of the three NS3 helicase domains. Saturation transfer difference nuclear magnetic resonance (NMR) experiments were additionally used to evaluate the binding strength of the fragments, revealing dissociation constants (KD) in the lower mM range. We conclude that the NS3 helicase of flaviviruses is a viable drug target. The data obtained open opportunities towards structure-based design of first-in-class anti-ZIKV compounds, as well as pan-flaviviral therapeutics.

Synthesis, characterization and crystal structures of novel fluorinated di(thiazolyl)benzene derivatives

2019 ◽  
Vol 6 (6) ◽  
pp. 780-790 ◽  
Author(s):  
Maciej Barłóg ◽  
Ihor Kulai ◽  
Xiaozhou Ji ◽  
Nattamai Bhuvanesh ◽  
Somnath Dey ◽  
...  
Keyword(s):  
Crystal Structures ◽  
Benzene Derivatives ◽  
Stille Coupling ◽  
X Ray ◽  
X Ray Crystallography ◽  
Microwave Assisted

A series of 11 novel fluorinated and non-fluorinated di(thiazolyl)benzenes have been synthesized via microwave assisted Stille coupling and characterized using X-ray crystallography.

scholarly journals Structure based protein engineering to confer selectivity of guanine deaminase

2014 ◽  
Vol 70 (a1) ◽  
pp. C437-C437
Author(s):  
Aruna Bitra ◽  
Ruchi Anand
Keyword(s):  
Crystal Structures ◽  
Active Site ◽  
Catalytic Mechanism ◽  
Catalytic Efficiency ◽  
Structural Basis ◽  
Active Site Residues ◽  
X Ray ◽  
Secondary Activity ◽  
Guanine Deaminase ◽  
Differential Selectivity

Guanine deaminases (GDs) are important enzymes involved in both purine metabolism and nucleotide anabolism pathways. Here we present the molecular and catalytic mechanism of NE0047 and use the information obtained to engineer specific enzyme activities. NE0047 from Nitrosomonas europaea was found to be a high fidelity guanine deaminase (catalytic efficiency of 1.2 × 105 M–1 s–1). However; it exhibited secondary activity towards the structurally non-analogous triazine based compound ammeline. The X-ray structure of NE0047 in the presence of the substrate analogue 8-azaguanine help establish that the enzyme exists as a biological dimer and both the proper closure of the C-terminal loop and cross talk via the dimeric interface is crucial for conferring catalytic activity. It was further ascertained that the highly conserved active site residues Glu79 and Glu143 facilitate the deamination reaction by serving as proton shuttles. Moreover, to understand the structural basis of dual substrate specificity, X-ray structures of NE0047 in complex with a series of nucleobase analogs, nucleosides and substrate ammeline were determined. The crystal structures demonstrated that any substitutions in the parent substrates results in the rearrangement of the ligand in a catalytically unfavorable orientation and also impede the closure of catalytically important loop, thereby abrogating activity. However, ammeline was able to adopt a catalytically favorable orientation which, also allowed for proper loop closure. Based on the above knowledge of the crystal structures and the catalytic mechanism, the active site was subsequently engineered to fine-tune NE0047 activity. The mutated versions of the enzyme were designed so that they can function either exclusively as a GD or serve as specific ammeline deaminases. For example, mutations in the active site E143D and N66A confer the enzyme to be an unambiguous GD with no secondary activity towards ammeline. On the other hand, the N66Q mutant of NE0047 only deaminates ammeline. Additionally, a series of crystal structures of the mutant versions were solved that shed light on the structural basis of this differential selectivity.

Synthesis and crystal structures of three novel benzimidazole/benzoindolizine hybrids

2016 ◽  
Vol 71 (3) ◽  
pp. 231-239 ◽  
Author(s):  
Roumaissa Belguedj ◽  
Sofiane Bouacida ◽  
Hocine Merazig ◽  
Ali Belfaitah ◽  
Aissa Chibani ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Nmr Spectroscopy ◽  
Crystal Structures ◽  
Dipolar Cycloaddition ◽  
Dimethyl Acetylenedicarboxylate ◽  
X Ray ◽  
X Ray Crystallography ◽  
Dimethyl Maleate

AbstractThree benzoindolizine derivatives, 1, 2, and 3, were obtained via 1,3-dipolar cycloaddition. The reaction of 1-(2′-benzimidazolylmethyl)isoquinolinium ylides with dimethyl acetylenedicarboxylate gave a mixture of pyrrolo[2,1-a]isoquinoline-1,2-dicarboxylate (1) and 1,10b-dihydropyrrolo[2,1-a]isoquinoline-1,2-dicarboxylate (2) derivatives containing a benzimidazole moiety. The reaction of this isoquinolinium N-ylide with dimethyl maleate gave an unexpected 2,3-dihydropyrrolo[2,1-a]isoquinoline-1,2-dicarboxylate (3). The structures of all reported compounds have been examined by X-ray crystallography, mass spectrometry, and NMR spectroscopy.

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