scholarly journals A Small Chemical Mimicking Actin Binding to Myosin and Putative Structural Changes of Myosin Molecule upon Ligand Binding

2014 ◽  
Vol 106 (2) ◽  
pp. 611a
Author(s):  
Takayuki Miyanishi ◽  
Io Omotuyi ◽  
Taku Yamaguchi
Keyword(s):  
Ligand Binding ◽  
Structural Changes ◽  
Actin Binding ◽  
Myosin Molecule ◽  
Small Chemical

scholarly journals Interbase-FRET binding assay for pre-microRNAs

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Mattias Bood ◽  
Anna Wypijewska del Nogal ◽  
Jesper R. Nilsson ◽  
Fredrik Edfeldt ◽  
Anders Dahlén ◽  
...  
Keyword(s):  
Ligand Binding ◽  
Drug Targets ◽  
Structural Changes ◽  
Small Sample Size ◽  
Resonance Energy ◽  
Small Sample ◽  
Titration Calorimetry ◽  
Drug Candidate ◽  
Binding Studies ◽  
Aberrant Expression

AbstractThe aberrant expression of microRNAs (miRs) has been linked to several human diseases. A promising approach for targeting these anomalies is the use of small-molecule inhibitors of miR biogenesis. These inhibitors have the potential to (i) dissect miR mechanisms of action, (ii) discover new drug targets, and (iii) function as new therapeutic agents. Here, we designed Förster resonance energy transfer (FRET)-labeled oligoribonucleotides of the precursor of the oncogenic miR-21 (pre-miR-21) and used them together with a set of aminoglycosides to develop an interbase-FRET assay to detect ligand binding to pre-miRs. Our interbase-FRET assay accurately reports structural changes of the RNA oligonucleotide induced by ligand binding. We demonstrate its application in a rapid, qualitative drug candidate screen by assessing the relative binding affinity between 12 aminoglycoside antibiotics and pre-miR-21. Surface plasmon resonance (SPR) and isothermal titration calorimetry (ITC) were used to validate our new FRET method, and the accuracy of our FRET assay was shown to be similar to the established techniques. With its advantages over SPR and ITC owing to its high sensitivity, small sample size, straightforward technique and the possibility for high-throughput expansion, we envision that our solution-based method can be applied in pre-miRNA–target binding studies.

scholarly journals Cofilin is a pH sensor for actin free barbed end formation: role of phosphoinositide binding

2008 ◽  
Vol 183 (5) ◽  
pp. 865-879 ◽  
Author(s):  
Christian Frantz ◽  
Gabriela Barreiro ◽  
Laura Dominguez ◽  
Xiaoming Chen ◽  
Robert Eddy ◽  
...  
Keyword(s):  
Actin Filament ◽  
Structural Changes ◽  
Ph Sensor ◽  
Ph Sensitive ◽  
Actin Binding ◽  
Filamentous Actin ◽  
Filament Dynamics ◽  
Ph Dependent ◽  
Dynamics Simulations

Newly generated actin free barbed ends at the front of motile cells provide sites for actin filament assembly driving membrane protrusion. Growth factors induce a rapid biphasic increase in actin free barbed ends, and we found both phases absent in fibroblasts lacking H+ efflux by the Na-H exchanger NHE1. The first phase is restored by expression of mutant cofilin-H133A but not unphosphorylated cofilin-S3A. Constant pH molecular dynamics simulations and nuclear magnetic resonance (NMR) reveal pH-sensitive structural changes in the cofilin C-terminal filamentous actin binding site dependent on His133. However, cofilin-H133A retains pH-sensitive changes in NMR spectra and severing activity in vitro, which suggests that it has a more complex behavior in cells. Cofilin activity is inhibited by phosphoinositide binding, and we found that phosphoinositide binding is pH-dependent for wild-type cofilin, with decreased binding at a higher pH. In contrast, phosphoinositide binding by cofilin-H133A is attenuated and pH insensitive. These data suggest a molecular mechanism whereby cofilin acts as a pH sensor to mediate a pH-dependent actin filament dynamics.

Growth-regulatory Human Galectin-1: Crystallographic Characterisation of the Structural Changes Induced by Single-site Mutations and their Impact on the Thermodynamics of Ligand Binding

2004 ◽  
Vol 343 (4) ◽  
pp. 957-970 ◽  
Author(s):  
María F. López-Lucendo ◽  
Dolores Solís ◽  
Sabine André ◽  
Jun Hirabayashi ◽  
Ken-ichi Kasai ◽  
...  
Keyword(s):  
Ligand Binding ◽  
Structural Changes ◽  
Single Site

Cytoglobin ligand binding regulated by changing haem-co-ordination in response to intramolecular disulfide bond formation and lipid interaction

2014 ◽  
Vol 465 (1) ◽  
pp. 127-137 ◽  
Author(s):  
Penny Beckerson ◽  
Michael T. Wilson ◽  
Dimitri A. Svistunenko ◽  
Brandon J. Reeder
Keyword(s):  
Ligand Binding ◽  
Redox State ◽  
Structural Changes ◽  
Bond Formation ◽  
Lipid Binding ◽  
Binding Kinetics ◽  
Disulfide Bond Formation ◽  
Physiological Properties ◽  
Intramolecular Disulfide Bond ◽  
Lipid Interaction

The redox state of the two-surface exposed cysteine residues in cytoglobin (Cygb) regulates the biochemical and potential physiological properties of the protein. Significant changes to ligand-binding kinetics, peroxidase activity and lipid-binding-induced structural changes are observed.

scholarly journals Evidence for a Conformational Change in Actin Induced by Fimbrin (N375) Binding

1997 ◽  
Vol 139 (2) ◽  
pp. 387-396 ◽  
Author(s):  
Dorit Hanein ◽  
Paul Matsudaira ◽  
David J. DeRosier
Keyword(s):  
Actin Filaments ◽  
Structural Changes ◽  
Three Dimensional ◽  
Binding Domain ◽  
Actin Binding ◽  
Dimensional Structure ◽  
Binding Surface ◽  
Actin Binding Domain ◽  
Actin Structure ◽  
Signal Transduction Proteins

Fimbrin belongs to a superfamily of actin cross-linking proteins that share a conserved 27-kD actin-binding domain. This domain contains a tandem duplication of a sequence that is homologous to calponin. Calponin homology (CH) domains not only cross-link actin filaments into bundles and networks, but they also bind intermediate filaments and some signal transduction proteins to the actin cytoskeleton. This fundamental role of CH domains as a widely used actin-binding domain underlines the necessity to understand their structural interaction with actin. Using electron cryomicroscopy, we have determined the three-dimensional structure of F-actin and F-actin decorated with the NH2-terminal CH domains of fimbrin (N375). In a difference map between actin filaments and N375-decorated actin, one end of N375 is bound to a concave surface formed between actin subdomains 1 and 2 on two neighboring actin monomers. In addition, a fit of the atomic model for the actin filament to the maps reveals the actin residues that line, the binding surface. The binding of N375 changes actin, which we interpret as a movement of subdomain 1 away from the bound N375. This change in actin structure may affect its affinity for other actin-binding proteins and may be part of the regulation of the cytoskeleton itself. Difference maps between actin and actin decorated with other proteins provides a way to look for novel structural changes in actin.

scholarly journals Cross-subunit Interactions that Stabilize Open States Mediate Gating in NMDA Receptors

2020 ◽  
Author(s):  
Gary J Iacobucci ◽  
Han Wen ◽  
Matthew B Helou ◽  
Wenjun Zheng ◽  
Gabriela K Popescu
Keyword(s):  
Ligand Binding ◽  
Nmda Receptors ◽  
Structural Changes ◽  
Chemical Interaction ◽  
Transmembrane Helix ◽  
Binding Domain ◽  
Ligand Binding Domain ◽  
Activation Reaction ◽  
Mechanical Coupling ◽  
Current Decay

ABSTRACTNMDA receptors are excitatory channels with critical functions in the physiology of central synapses. Their activation reaction proceeds as a series of kinetically distinguishable, reversible steps, whose structural bases are of current interest. Very likely, the earliest steps in the activation reaction include glutamate binding to and compression of the ligand-binding domain. Later, three short linkers transduce this movement to open the gate by mechanical coupling with transmembrane helices. Here, we used double-mutant cycle analyses to demonstrate that a direct chemical interaction between GluN1-I642 (on M3) and GluN2A-L550 (on L1-M1) stabilizes receptors after they have opened, and therefore represents one of the structural changes that occur late in the activation reaction. This native interaction extends the current decay, and its absence predicts deficits in charge transfer by GluN1-I642L, a pathogenic human variant.SIGNIFICANCE STATEMENTNMDA receptors are glutamatergic channels whose activations control the strength of excitatory synapses in the central nervous system. Agonist binding initiates a complex activation reaction that consists of a stepwise sequence of reversible isomerizations. In addition to previously identified steps in this series, which include agonist-induced closure of the ligand-binding lobes, and the subsequent mechanical pulling by the ligand-binding domain on the gate-forming transmembrane helix, we identify a new cross-subunit interaction, which stabilizes open receptors and slows the rate of the current decay. Naturally occurring NMDA receptor variants lacking this interaction are pathogenic.

scholarly journals Capturing Peptide–GPCR Interactions and Their Dynamics

Molecules ◽  
2020 ◽  
Vol 25 (20) ◽  
pp. 4724
Author(s):  
Anette Kaiser ◽  
Irene Coin
Keyword(s):  
Ligand Binding ◽  
Conformational Changes ◽  
Structural Changes ◽  
Structural Heterogeneity ◽  
Receptor Interaction ◽  
Spectroscopic Techniques ◽  
Peptide Receptors ◽  
Physiological Processes ◽  
High Dynamics ◽  
G Protein Coupled

Many biological functions of peptides are mediated through G protein-coupled receptors (GPCRs). Upon ligand binding, GPCRs undergo conformational changes that facilitate the binding and activation of multiple effectors. GPCRs regulate nearly all physiological processes and are a favorite pharmacological target. In particular, drugs are sought after that elicit the recruitment of selected effectors only (biased ligands). Understanding how ligands bind to GPCRs and which conformational changes they induce is a fundamental step toward the development of more efficient and specific drugs. Moreover, it is emerging that the dynamic of the ligand–receptor interaction contributes to the specificity of both ligand recognition and effector recruitment, an aspect that is missing in structural snapshots from crystallography. We describe here biochemical and biophysical techniques to address ligand–receptor interactions in their structural and dynamic aspects, which include mutagenesis, crosslinking, spectroscopic techniques, and mass-spectrometry profiling. With a main focus on peptide receptors, we present methods to unveil the ligand–receptor contact interface and methods that address conformational changes both in the ligand and the GPCR. The presented studies highlight a wide structural heterogeneity among peptide receptors, reveal distinct structural changes occurring during ligand binding and a surprisingly high dynamics of the ligand–GPCR complexes.

scholarly journals Tumor Suppressor Activity of Profilin Requires a Functional Actin Binding Site

2004 ◽  
Vol 15 (4) ◽  
pp. 1600-1608 ◽  
Author(s):  
Nina Wittenmayer ◽  
Burkhard Jandrig ◽  
Martin Rothkegel ◽  
Kathrin Schlüter ◽  
Wolfgang Arnold ◽  
...  
Keyword(s):  
Tumor Suppressor ◽  
Ligand Binding ◽  
Binding Site ◽  
Nude Mice ◽  
Human Cancer ◽  
Cancer Cell Line ◽  
Actin Binding ◽  
Suppressor Activity ◽  
Tumor Suppressor Activity ◽  
Actin Binding Site

Profilin 1 (PFN1) is a regulator of the microfilament system and is involved in various signaling pathways. It interacts with many cytoplasmic and nuclear ligands. The importance of PFN1 for human tissue differentiation has been demonstrated by the findings that human cancer cells, expressing conspicuously low PFN1 levels, adopt a nontumorigenic phenotype upon raising their PFN1 level. In the present study, we characterize the ligand binding site crucial for profilin's tumor suppressor activity. Starting with CAL51, a human breast cancer cell line highly tumorigenic in nude mice, we established stable clones that express PFN1 mutants differentially defective in ligand binding. Clones expressing PFN1 mutants with reduced binding to either poly-proline-stretch ligands or phosphatidyl-inositol-4,5-bisphosphate, but with a functional actin binding site, were normal in growth, adhesion, and anchorage dependence, with only a weak tendency to elicit tumors in nude mice, similar to controls expressing wild-type PFN1. In contrast, clones expressing a mutant with severely reduced capacity to bind actin still behaved like the parental CAL51 and were highly tumorigenic. We conclude that the actin binding site on profilin is instrumental for normal differentiation of human epithelia and the tumor suppressor function of PFN1.

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