scholarly journals Structural basis for biomolecular recognition in overlapping binding sites in a diiron enzyme system

2014 ◽  
Vol 5 (1) ◽  
Author(s):  
Justin F. Acheson ◽  
Lucas J. Bailey ◽  
Nathaniel L. Elsen ◽  
Brian G. Fox
Keyword(s):  
Binding Sites ◽  
Enzyme System ◽  
Structural Basis ◽  
Biomolecular Recognition ◽  
Diiron Enzyme

scholarly journals Local frustration determines loop opening during the catalytic cycle of an oxidoreductase

2019 ◽  
Author(s):  
Lukas L. Stelzl ◽  
Despoina A.I. Mavridou ◽  
Emmanuel Saridakis ◽  
Diego Gonzalez ◽  
Andrew J. Baldwin ◽  
...  
Keyword(s):  
Active Site ◽  
Binding Sites ◽  
Protein Function ◽  
Catalytic Cycle ◽  
Biomolecular Recognition ◽  
Competing Interactions ◽  
Protein Binding Sites ◽  
Loop Region ◽  
Cysteine Thiols ◽  
Small Chemical

AbstractLocal structural frustration, the existence of mutually exclusive competing interactions, may explain why some proteins are dynamic while others are rigid. Frustration is thought to underpin biomolecular recognition and the flexibility of protein binding sites. Here we show how a small chemical modification, the oxidation of two cysteine thiols to a disulfide bond, during the catalytic cycle of the N-terminal domain of the key bacterial oxidoreductase DsbD (nDsbD), introduces frustration ultimately influencing protein function. In oxidized nDsbD, local frustration disrupts the packing of the protective cap-loop region against the active site allowing loop opening. By contrast, in reduced nDsbD the cap loop is rigid, always protecting the active-site thiols from the oxidizing environment of the periplasm. Our results point towards an intricate coupling between the dynamics of the active-site cysteines and of the cap loop which modulates the association reactions of nDsbD with its partners resulting in optimized protein function.

scholarly journals Cryo-EM structure of the complete and ligand-saturated insulin receptor ectodomain

2019 ◽  
Author(s):  
Theresia Gutmann ◽  
Ingmar Schäfer ◽  
Chetan Poojari ◽  
Beate Brankatschk ◽  
Ilpo Vattulainen ◽  
...  
Keyword(s):  
Insulin Receptor ◽  
Binding Sites ◽  
Structural Model ◽  
Receptor Site ◽  
Insulin Binding ◽  
Proximal Region ◽  
Dynamics Simulation ◽  
Structural Basis ◽  
Structure Based Drug Design ◽  
Receptor Interactions

AbstractGlucose homeostasis and growth essentially depend on the peptide hormone insulin engaging its receptor. Despite biochemical and structural advances, a fundamental contradiction has persisted in the current understanding of insulin ligand–receptor interactions. While biochemistry predicts two distinct insulin binding sites, 1 and 2, recent structural analyses have only resolved site 1. Using a combined approach of cryo-EM and atomistic molecular dynamics simulation, we determined the structure of the entire dimeric insulin receptor ectodomain saturated with four insulin molecules. Complementing the previously described insulin–site 1 interaction, we present the first view of insulin bound to the discrete insulin receptor site 2. Insulin binding stabilizes the receptor ectodomain in a T-shaped conformation wherein the membrane-proximal domains converge and contact each other. These findings expand the current models of insulin binding to its receptor and of its regulation. In summary, we provide the structural basis enabling a comprehensive description of ligand–receptor interactions that ultimately will inform new approaches to structure-based drug design.In briefA cryo-EM structure of the complete insulin receptor ectodomain saturated with four insulin ligands is reported. The structural model of the insulin–insulin receptor complex adopts a T-shaped conformation, reveals two additional insulin-binding sites potentially involved in the initial interaction of insulin with its receptor, and resolves the membrane proximal region.

Structural characterization of amorphous calcium carbonate-binding protein: an insight into the mechanism of amorphous calcium carbonate formation

2013 ◽  
Vol 453 (2) ◽  
pp. 179-186 ◽  
Author(s):  
Jingtan Su ◽  
Xiao Liang ◽  
Qiang Zhou ◽  
Guiyou Zhang ◽  
Hongzhong Wang ◽  
...  
Keyword(s):  
Calcium Carbonate ◽  
Binding Sites ◽  
Binding Protein ◽  
Homology Modelling ◽  
Size Exclusion ◽  
Structural Basis ◽  
Amorphous Calcium Carbonate ◽  
Carbonate Formation

ACC (amorphous calcium carbonate) plays an important role in biomineralization process for its function as a precursor for calcium carbonate biominerals. However, it is unclear how biomacromolecules regulate the formation of ACC precursor in vivo. In the present study, we used biochemical experiments coupled with bioinformatics approaches to explore the mechanisms of ACC formation controlled by ACCBP (ACC-binding protein). Size-exclusion chromatography, chemical cross-linking experiments and negative staining electron microscopy reveal that ACCBP is a decamer composed of two adjacent pentamers. Sequence analyses and fluorescence quenching results indicate that ACCBP contains two Ca2+-binding sites. The results of in vitro crystallization experiments suggest that one Ca2+-binding site is critical for ACC formation and the other site affects the ACC induction efficiency. Homology modelling demonstrates that the Ca2+-binding sites of pentameric ACCBP are arranged in a 5-fold symmetry, which is the structural basis for ACC formation. To the best of our knowledge, this is the first report on the structural basis for protein-induced ACC formation and it will significantly improve our understanding of the amorphous precursor pathway.

scholarly journals Cryo-EM structures of a lipid-sensitive pentameric ligand-gated ion channel embedded in a phosphatidylcholine-only bilayer

2020 ◽  
Vol 117 (3) ◽  
pp. 1788-1798 ◽  
Author(s):  
Pramod Kumar ◽  
Yuhang Wang ◽  
Zhening Zhang ◽  
Zhiyu Zhao ◽  
Gisela D. Cymes ◽  
...  
Keyword(s):  
Ion Channel ◽  
Binding Sites ◽  
Electric Organ ◽  
Structural Models ◽  
Erwinia Chrysanthemi ◽  
Structural Basis ◽  
Agonist Binding ◽  
Nicotinic Acetylcholine ◽  
Bacterial Homolog ◽  
Straightforward Interpretation

The lipid dependence of the nicotinic acetylcholine receptor from the Torpedo electric organ has long been recognized, and one of the most consistent experimental observations is that, when reconstituted in membranes formed by zwitterionic phospholipids alone, exposure to agonist fails to elicit ion-flux activity. More recently, it has been suggested that the bacterial homolog ELIC (Erwinia chrysanthemi ligand-gated ion channel) has a similar lipid sensitivity. As a first step toward the elucidation of the structural basis of this phenomenon, we solved the structures of ELIC embedded in palmitoyl-oleoyl-phosphatidylcholine- (POPC-) only nanodiscs in both the unliganded (4.1-Å resolution) and agonist-bound (3.3 Å) states using single-particle cryoelectron microscopy. Comparison of the two structural models revealed that the largest differences occur at the level of loop C—at the agonist-binding sites—and the loops at the interface between the extracellular and transmembrane domains (ECD and TMD, respectively). On the other hand, the transmembrane pore is occluded in a remarkably similar manner in both structures. A straightforward interpretation of these findings is that POPC-only membranes frustrate the ECD–TMD coupling in such a way that the “conformational wave” of liganded-receptor gating takes place in the ECD and the interfacial M2–M3 linker but fails to penetrate the membrane and propagate into the TMD. Furthermore, analysis of the structural models and molecular simulations suggested that the higher affinity for agonists characteristic of the open- and desensitized-channel conformations results, at least in part, from the tighter confinement of the ligand to its binding site; this limits the ligand’s fluctuations, and thus delays its escape into bulk solvent.

scholarly journals Analysis of the structural requirements of sugar binding to the liver, brain and insulin-responsive glucose transporters expressed in oocytes

1993 ◽  
Vol 294 (3) ◽  
pp. 753-760 ◽  
Author(s):  
C A Colville ◽  
M J Seatter ◽  
G W Gould
Keyword(s):  
Hydrogen Bond ◽  
Binding Sites ◽  
Glucose Transporters ◽  
Binding Pocket ◽  
Structural Basis ◽  
Sugar Binding ◽  
Glucose Analogues ◽  
Sugar Recognition

We have expressed the liver (GLUT 2), brain (GLUT 3) and insulin-responsive (GLUT 4) glucose transporters in oocytes from Xenopus laevis by microinjection of in vitro-transcribed mRNA. Using a range of halogeno- and deoxy-glucose analogues, and other hexoses, we have studied the structural basis of sugar binding to these different isoforms. We show that a hydrogen bond to the C-3 position is involved in sugar binding for all three isoforms, but that the direction of this hydrogen bond is different in GLUT 2 from either GLUT 1, 3 or 4. Hydrogen-bonding at the C-4 position is also involved in sugar recognition by all three isoforms, but we propose that in GLUT 3 this hydrogen bond plays a less significant role than in GLUT 2 and 4. In all transporters we propose that the C-4 position is directed out of the sugar-binding pocket. The role of the C-6 position is also discussed. In addition, we have analysed the ability of fructopyranose and fructofuranose analogues to inhibit the transport mediated by GLUT2. We show that fructofuranose analogues, but not fructopyranose analogues, are efficient inhibitors of transport mediated by GLUT 2, and therefore suggest that GLUT 2 accommodates D-glucose as a pyranose ring, but D-fructose as a furanose ring. Models for the binding sites of GLUT 2, 3 and 4 are presented.

scholarly journals The structural basis for inhibition of ribosomal translocation by viomycin

2020 ◽  
Vol 117 (19) ◽  
pp. 10271-10277
Author(s):  
Ling Zhang ◽  
Ying-Hui Wang ◽  
Xing Zhang ◽  
Laura Lancaster ◽  
Jie Zhou ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Binding Sites ◽  
Ribosomal Subunit ◽  
Structural Basis ◽  
23S Rrna ◽  
Subunit Dissociation ◽  
Classical State ◽  
State Transfer ◽  
70S Ribosome ◽  
Intersubunit Rotation

Viomycin, an antibiotic that has been used to fight tuberculosis infections, is believed to block the translocation step of protein synthesis by inhibiting ribosomal subunit dissociation and trapping the ribosome in an intermediate state of intersubunit rotation. The mechanism by which viomycin stabilizes this state remains unexplained. To address this, we have determined cryo-EM and X-ray crystal structures of Escherichia coli 70S ribosome complexes trapped in a rotated state by viomycin. The 3.8-Å resolution cryo-EM structure reveals a ribosome trapped in the hybrid state with 8.6° intersubunit rotation and 5.3° rotation of the 30S subunit head domain, bearing a single P/E state transfer RNA (tRNA). We identify five different binding sites for viomycin, four of which have not been previously described. To resolve the details of their binding interactions, we solved the 3.1-Å crystal structure of a viomycin-bound ribosome complex, revealing that all five viomycins bind to ribosomal RNA. One of these (Vio1) corresponds to the single viomycin that was previously identified in a complex with a nonrotated classical-state ribosome. Three of the newly observed binding sites (Vio3, Vio4, and Vio5) are clustered at intersubunit bridges, consistent with the ability of viomycin to inhibit subunit dissociation. We propose that one or more of these same three viomycins induce intersubunit rotation by selectively binding the rotated state of the ribosome at dynamic elements of 16S and 23S rRNA, thus, blocking conformational changes associated with molecular movements that are required for translocation.

Structural basis of carbohydrate binding in domain C of a type I pullulanase from Paenibacillus barengoltzii

2020 ◽  
Vol 76 (5) ◽  
pp. 447-457
Author(s):  
Ping Huang ◽  
Shiwang Wu ◽  
Shaoqing Yang ◽  
Qiaojuan Yan ◽  
Zhengqiang Jiang
Keyword(s):  
Binding Site ◽  
Binding Sites ◽  
Structural Basis ◽  
Carbohydrate Binding ◽  
Type I ◽  
Debranching Enzyme ◽  
Crystal Forms ◽  
Aromatic Residues ◽  
Starch Debranching Enzyme ◽  
Paenibacillus Barengoltzii

Pullulanase (EC 3.2.1.41) is a well known starch-debranching enzyme that catalyzes the cleavage of α-1,6-glycosidic linkages in α-glucans such as starch and pullulan. Crystal structures of a type I pullulanase from Paenibacillus barengoltzii (PbPulA) and of PbPulA in complex with maltopentaose (G5), maltohexaose (G6)/α-cyclodextrin (α-CD) and β-cyclodextrin (β-CD) were determined in order to better understand substrate binding to this enzyme. PbPulA belongs to glycoside hydrolase (GH) family 13 subfamily 14 and is composed of three domains (CBM48, A and C). Three carbohydrate-binding sites identified in PbPulA were located in CBM48, near the active site and in domain C, respectively. The binding site in CBM48 was specific for β-CD, while that in domain C has not been reported for other pullulanases. The domain C binding site had higher affinity for α-CD than for G6; a small motif (FGGEH) seemed to be one of the major determinants for carbohydrate binding in this domain. Structure-based mutations of several surface-exposed aromatic residues in CBM48 and domain C had a debilitating effect on the activity of the enzyme. These results suggest that both CBM48 and domain C play a role in binding substrates. The crystal forms described contribute to the understanding of pullulanase domain–carbohydrate interactions.

scholarly journals Structural basis of ion transport and inhibition in ferroportin

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Yaping Pan ◽  
Zhenning Ren ◽  
Shuai Gao ◽  
Jiemin Shen ◽  
Lie Wang ◽  
...  
Keyword(s):  
Binding Sites ◽  
Metal Ion ◽  
Peptide Hormone ◽  
Structural Basis ◽  
Deficiency Anemia ◽  
Pharmacological Intervention ◽  
Ion Binding ◽  
Coupled Transport ◽  
Metal Ion Binding ◽  
Ion Binding Sites

Abstract Ferroportin is an iron exporter essential for releasing cellular iron into circulation. Ferroportin is inhibited by a peptide hormone, hepcidin. In humans, mutations in ferroportin lead to ferroportin diseases that are often associated with accumulation of iron in macrophages and symptoms of iron deficiency anemia. Here we present the structures of the ferroportin from the primate Philippine tarsier (TsFpn) in the presence and absence of hepcidin solved by cryo-electron microscopy. TsFpn is composed of two domains resembling a clamshell and the structure defines two metal ion binding sites, one in each domain. Both structures are in an outward-facing conformation, and hepcidin binds between the two domains and reaches one of the ion binding sites. Functional studies show that TsFpn is an electroneutral H+/Fe2+ antiporter so that transport of each Fe2+ is coupled to transport of two H+ in the opposite direction. Perturbing either of the ion binding sites compromises the coupled transport of H+ and Fe2+. These results establish the structural basis of metal ion binding, transport and inhibition in ferroportin and provide a blueprint for targeting ferroportin in pharmacological intervention of ferroportin diseases.

scholarly journals Cation-π bonding and amino-aromatic interactions in the biomolecular recognition of substituted ammonium ligands

1996 ◽  
Vol 319 (1) ◽  
pp. 1-8 ◽  
Author(s):  
Nigel S. SCRUTTON ◽  
Andrew R.C. RAINE
Keyword(s):  
Protein Interactions ◽  
Binding Sites ◽  
Biologically Active ◽  
Biomolecular Recognition ◽  
Structure Based Drug Design ◽  
Protein Architecture ◽  
Aromatic Interactions ◽  
Ligand Binding Sites ◽  
Amine Ligands

Cation-π bonds and amino-aromatic interactions are known to be important contributors to protein architecture and stability, and their role in ligand-protein interactions has also been reported. Many biologically active amines contain substituted ammonium moieties, and cation-π bonding and amino-aromatic interactions often enable these molecules to associate with proteins. The role of organic cation-π bonding and amino-aromatic interactions in the recognition of small-molecule amines and peptides by proteins is an important topic for those involved in structure-based drug design, and although the number of structures determined for proteins displaying these interactions is small, general features are beginning to emerge. This review explores the role of cation-π bonding and amino-aromatic interactions in the biological molecular recognition of amine ligands. Perspectives on the design of ammonium-ligand-binding sites are also discussed.

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