scholarly journals Prediction and Validation of a Druggable Site on Virulence Factor of Drug Resistant Burkholderia Cenocepacia

2020 ◽  
Author(s):  
Kanhaya Lal ◽  
Rafael Bermo ◽  
Jonathan Cramer ◽  
Francesca Vasile ◽  
Beat Ernst ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Virtual Screening ◽  
Binding Site ◽  
Virulence Factor ◽  
Opportunistic Pathogen ◽  
Binding Pocket ◽  
Burkholderia Cenocepacia ◽  
Drug Resistant ◽  
Novel Approach ◽  
Virtual Screening Approach

<br><p> </p> <p>We use a virtual screening approach to explore the binding pocket close to the fucose binding site in a lectin from opportunistic pathogen. This is a novel approach for designing anti-adhesive drugs, and it has been very successful since we were able to obtain leads with sub millimolar affinity. Furthermore, the crystal structure of the target protein complexed with the fragment validated the existence of this secondary binding site, opening route for new design of inhibitors</p>

scholarly journals Prediction and Validation of a Druggable Site on Virulence Factor of Drug Resistant Burkholderia Cenocepacia

2020 ◽  
Author(s):  
Kanhaya Lal ◽  
Rafael Bermo ◽  
Jonathan Cramer ◽  
Francesca Vasile ◽  
Beat Ernst ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Virtual Screening ◽  
Binding Site ◽  
Virulence Factor ◽  
Opportunistic Pathogen ◽  
Binding Pocket ◽  
Burkholderia Cenocepacia ◽  
Drug Resistant ◽  
Novel Approach ◽  
Virtual Screening Approach

<br><p> </p> <p>We use a virtual screening approach to explore the binding pocket close to the fucose binding site in a lectin from opportunistic pathogen. This is a novel approach for designing anti-adhesive drugs, and it has been very successful since we were able to obtain leads with sub millimolar affinity. Furthermore, the crystal structure of the target protein complexed with the fragment validated the existence of this secondary binding site, opening route for new design of inhibitors</p>

scholarly journals Prediction and Validation of a Druggable Site on Virulence Factor of Drug Resistant Burkholderia cenocepacia

2021 ◽  
Author(s):  
Kanhaya Lal ◽  
Rafael Bermeo ◽  
Jonathan Cramer ◽  
Francesca Vasile ◽  
Beat Ernst ◽  
...  
Keyword(s):  
Virulence Factor ◽  
Burkholderia Cenocepacia ◽  
Drug Resistant

scholarly journals Characterization of the Acetate Binding Pocket in the Methanosarcina thermophila Acetate Kinase

2005 ◽  
Vol 187 (7) ◽  
pp. 2386-2394 ◽  
Author(s):  
Cheryl Ingram-Smith ◽  
Andrea Gorrell ◽  
Sarah H. Lawrence ◽  
Prabha Iyer ◽  
Kerry Smith ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Binding Site ◽  
Binding Pocket ◽  
Acetate Kinase ◽  
Phosphoryl Group ◽  
Acetyl Phosphate ◽  
Hydrophobic Pocket ◽  
Methanosarcina Thermophila ◽  
Methyl Group ◽  
Substrate Binding Sites

ABSTRACT Acetate kinase catalyzes the reversible magnesium-dependent synthesis of acetyl phosphate by transfer of the ATP γ-phosphoryl group to acetate. Inspection of the crystal structure of the Methanosarcina thermophila enzyme containing only ADP revealed a solvent-accessible hydrophobic pocket formed by residues Val93, Leu122, Phe179, and Pro232 in the active site cleft, which identified a potential acetate binding site. The hypothesis that this was a binding site was further supported by alignment of all acetate kinase sequences available from databases, which showed strict conservation of all four residues, and the recent crystal structure of the M. thermophila enzyme with acetate bound in this pocket. Replacement of each residue in the pocket produced variants with Km values for acetate that were 7- to 26-fold greater than that of the wild type, and perturbations of this binding pocket also altered the specificity for longer-chain carboxylic acids and acetyl phosphate. The kinetic analyses of variants combined with structural modeling indicated that the pocket has roles in binding the methyl group of acetate, influencing substrate specificity, and orienting the carboxyl group. The kinetic analyses also indicated that binding of acetyl phosphate is more dependent on interactions of the phosphate group with an unidentified residue than on interactions between the methyl group and the hydrophobic pocket. The analyses also indicated that Phe179 is essential for catalysis, possibly for domain closure. Alignments of acetate kinase, propionate kinase, and butyrate kinase sequences obtained from databases suggested that these enzymes have similar catalytic mechanisms and carboxylic acid substrate binding sites.

scholarly journals Crystal structure of a human neuronal nAChR extracellular domain in pentameric assembly: Ligand-bound α2 homopentamer

2016 ◽  
Vol 113 (34) ◽  
pp. 9635-9640 ◽  
Author(s):  
Nikolaos Kouvatsos ◽  
Petros Giastas ◽  
Dafni Chroni-Tzartou ◽  
Cornelia Poulopoulou ◽  
Socrates J. Tzartos
Keyword(s):  
Crystal Structure ◽  
Binding Site ◽  
Structural Information ◽  
Extracellular Domain ◽  
Binding Pocket ◽  
Nachr Subunit ◽  
Homologous Proteins ◽  
High Sequence Identity ◽  
Neuronal Nachr ◽  
Low Sensitivity

In this study we report the X-ray crystal structure of the extracellular domain (ECD) of the human neuronal α2 nicotinic acetylcholine receptor (nAChR) subunit in complex with the agonist epibatidine at 3.2 Å. Interestingly, α2 was crystallized as a pentamer, revealing the intersubunit interactions in a wild type neuronal nAChR ECD and the full ligand binding pocket conferred by two adjacent α subunits. The pentameric assembly presents the conserved structural scaffold observed in homologous proteins, as well as distinctive features, providing unique structural information of the binding site between principal and complementary faces. Structure-guided mutagenesis and electrophysiological data confirmed the presence of the α2(+)/α2(−) binding site on the heteromeric low sensitivity α2β2 nAChR and validated the functional importance of specific residues in α2 and β2 nAChR subunits. Given the pathological importance of the α2 nAChR subunit and the high sequence identity with α4 (78%) and other neuronal nAChR subunits, our findings offer valuable information for modeling several nAChRs and ultimately for structure-based design of subtype specific drugs against the nAChR associated diseases.

scholarly journals Identification of the site of oxidase substrate binding inScytalidium thermophilumcatalase

2018 ◽  
Vol 74 (10) ◽  
pp. 979-985 ◽  
Author(s):  
Yonca Yuzugullu Karakus ◽  
Gunce Goc ◽  
Sinem Balci ◽  
Briony A. Yorke ◽  
Chi H. Trinh ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Kinetic Analysis ◽  
Binding Site ◽  
Active Site ◽  
Oxidase Activity ◽  
Substrate Binding ◽  
Binding Pocket ◽  
Scytalidium Thermophilum ◽  
Lateral Channel

The catalase fromScytalidium thermophilumis a homotetramer containing a hemedin each active site. Although the enzyme has a classical monofunctional catalase fold, it also possesses oxidase activity towards a number of small organics, including catechol and phenol. In order to further investigate this, the crystal structure of the complex of the catalase with the classical catalase inhibitor 3-amino-1,2,4-triazole (3TR) was determined at 1.95 Å resolution. Surprisingly, no binding to the heme site was observed; instead, 3TR occupies a binding site corresponding to the NADPH-binding pocket in mammalian catalases at the entrance to a lateral channel leading to the heme. Kinetic analysis of site-directed mutants supports the assignment of this pocket as the binding site for oxidase substrates.

Critical residues for ligand binding in blade 2 of the propeller domain of the integrin αIIb subunit

2004 ◽  
Vol 91 (01) ◽  
pp. 111-118 ◽  
Author(s):  
Tatsushiro Tamura ◽  
Jun Yamanouchi ◽  
Shigeru Fujita ◽  
Takaaki Hato
Keyword(s):  
Crystal Structure ◽  
Ligand Binding ◽  
Binding Site ◽  
Thrombus Formation ◽  
Direct Interaction ◽  
Binding Pocket ◽  
Crystal Structure Analysis ◽  
Site Directed Mutagenesis ◽  
Structural Basis ◽  
Critical Residues

SummaryLigand binding to integrin αIIbβ3 is a key event of thrombus formation. The propeller domain of the αIIb subunit has been implicated in ligand binding. Recently, the ligand binding site of the αV propeller was determined by crystal structure analysis. However, the structural basis of ligand recognition by the αIIb propeller remains to be determined. In this study, we conducted site-directed mutagenesis of all residues located in the loops extending above blades 2 and 4 of the αIIb propeller, which are spatially close to, but distinct from, the loops that contain the binding site for an RGD ligand in the crystal structure of the αV propeller. Replacement by alanine of Q111, H112 or N114 in the loop within the blade 2 (the W2:2-3 loop in the propeller model) abolished binding of a ligand-mimetic antibody and fibrinogen to αIIbβ3 induced by different types of integrin activation including activation of αIIbβ3 by β3 cytoplasmic mutation. CHO cells stably expressing recombinant αIIbβ3 bearing Q111A, H112A or N114A mutation did not exhibit αIIbβ3mediated adhesion to fibrinogen. According to the crystal structure of αVβ3, the αV residue corresponding to αIIbN114 is exposed on the integrin surface and close to the RGD binding site. These results suggest that the Q111, H112 and N114 residues in the loop within blade 2 of the αIIb propeller are critical for ligand binding, possibly because of direct interaction with ligands or modulation of the RGD binding pocket.

Study of AMP-Activated Protein Kinase Agonists by Structure-Based Drug Designing

2009 ◽  
Vol 79-82 ◽  
pp. 2187-2190 ◽  
Author(s):  
Yea Huey Chang ◽  
Tin Yun Ho ◽  
Chieh Hsi Wu ◽  
Chien Yu Chen ◽  
Hung Jin Huang ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Crystal Structure ◽  
Protein Kinase ◽  
Virtual Screening ◽  
Homology Modeling ◽  
Binding Site ◽  
X Ray ◽  
Drug Designing ◽  
Key Residues ◽  
Amp Activated Protein Kinase

AMP-activated protein kinase (AMPK) is a metabolite- sensed protein kinase in various eukaryotes. The activated AMPK regulates important proteins which cause diabetes, obesity, metabolic aberrant, and also breast cancer. In this study, the yeast AMPK structure was used as a template to model the human AMPK structure. By homology modeling, the reliable AMPK structure was built, and the active binding site was defined corresponding to X-ray crystal structure of yeast AMPK By virtual screening the database., All the potent ligands had the H-bond interaction in the key residues, same as the control. Thus, we suggested the phenylamide derivates might be the potent AMPK agonists.

scholarly journals Crystal structure of CotA laccase complexed with 2,2-azinobis-(3-ethylbenzothiazoline-6-sulfonate) at a novel binding site

2016 ◽  
Vol 72 (4) ◽  
pp. 328-335 ◽  
Author(s):  
Zhongchuan Liu ◽  
Tian Xie ◽  
Qiuping Zhong ◽  
Ganggang Wang
Keyword(s):  
Crystal Structure ◽  
Binding Site ◽  
Binding Pocket ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
The Novel ◽  
Wild Type ◽  
Outer Coat ◽  
Cota Laccase ◽  
Key Residues

The CotA laccase fromBacillus subtilisis an abundant component of the spore outer coat and has been characterized as a typical laccase. The crystal structure of CotA complexed with 2,2-azinobis-(3-ethylbenzothiazoline-6-sulfonate) (ABTS) in a hole motif has been solved. The novel binding site was about 26 Å away from the T1 binding pocket. Comparison with known structures of other laccases revealed that the hole is a specific feature of CotA. The key residues Arg476 and Ser360 were directly bound to ABTS. Site-directed mutagenesis studies revealed that the residues Arg146, Arg429 and Arg476, which are located at the bottom of the novel binding site, are essential for the oxidation of ABTS and syringaldazine. Specially, a Thr480Phe variant was identified to be almost 3.5 times more specific for ABTS than for syringaldazine compared with the wild type. These results suggest this novel binding site for ABTS could be a potential target for protein engineering of CotA laccases.

scholarly journals A closed-homodimer structure of human importin-α1

2014 ◽  
Vol 70 (a1) ◽  
pp. C498-C498
Author(s):  
Hideyuki Miyatake ◽  
Akira Sanjoh ◽  
Go Matsuda ◽  
Yuko Tatsumi ◽  
Yoichi Miyamoto ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Binding Site ◽  
Binding Affinity ◽  
Binding Sites ◽  
Binding Pocket ◽  
Dimer Formation ◽  
Functional Aspects ◽  
Physicochemical Studies

We determined the crystal structure of N-terminal importin-β-binding (IBB) domain truncated human importin-α1 (ΔIBB-importin-α1) at 2.63Å resolution. The crystal structure of ΔIBB-importin-α1 is a novel closed-homodimer. The homodimer exists in an autoinhibition state in which both of the major and minor NLS-binding sites are completely buried in the homodimerization interface to avoid NLS binding. Importin-α1 is in dimer-monomer equilibration in solution. In the dimerization sate, the P1'-binding pocket in the minor NLS binding site plays a role to stabilize the dimer formation. The external K108 binds into the P1'-binding pocket that results in the autoinhibition of the NLS binding. The present closed-homodimer of ΔIBB-importin-α1 conjured the functional aspects of multimerization of importin-α1. The further physicochemical studies using full- and ΔIBB- importin-α1 reveal that the IBB domain is involved in the monomer-dimer equilibration; thereby the NLS binding affinity is kept even in the higher concentration of importin-α1. Owing to the multimerization property, importin-αs can autoinhibition the NLS binding, that may result in a variety of NLS recognition way.

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