scholarly journals The proximal and distal pockets of the H93G myoglobin cavity mutant bind identical ligands with different affinities: Quantitative analysis of imidazole and pyridine binding

2008 ◽  
Vol 22 (2-3) ◽  
pp. 123-141 ◽  
Author(s):  
Jing Du ◽  
Masanori Sono ◽  
John H. Dawson
Keyword(s):  
Ligand Binding ◽  
Magnetic Circular Dichroism ◽  
Sperm Whale ◽  
Heme Iron ◽  
Model Systems ◽  
Binding Affinities ◽  
Quantitative Evidence ◽  
High Affinity Binding Site ◽  
Free Heme ◽  
Exogenous Ligand

His93Gly sperm whale myoglobin (H93G Mb) has the proximal histidine ligand removed to create a cavity for exogenous ligand binding, making it a versatile template for the preparation of model heme complexes. In this study, we have measured the first and second ligand binding affinities of imidazole and pyridine to form mono- and bis-ligated ferric and ferrous H93G Mb complexes. Electronic absorption spectroscopy has been utilized to determine the binding affinities for the proximal (Kd1, first ligand) and distal (Kd2, second ligand) pockets of H93G Mb. Magnetic circular dichroism spectroscopy has been used to confirm the identity of the complexes. The binding affinities for the first ligand are one hundred- to one thousand-fold higher than those for the second ligand (Kd1«Kd2) for the same exogenous ligand. This is entirely opposite to what is seen with free heme in organic solvents whereKd1»Kd2. Thus, the proximal pocket is the high affinity binding site. The lower affinity for the distal pocket can be attributed to steric hindrance from the distal histidine. This report provides quantitative evidence for differential ligand binding affinities of the proximal and distal pockets of H93G Mb, a unique property that facilitates generation of heme iron derivatives not easily prepared with other heme model systems.

Ferric His93Gly myoglobin cavity mutant and its complexes with thioether and selenolate as heme protein models

2011 ◽  
Vol 15 (01) ◽  
pp. 29-38 ◽  
Author(s):  
Jing Du ◽  
Masanori Sono ◽  
John H. Dawson
Keyword(s):  
Active Sites ◽  
Magnetic Circular Dichroism ◽  
Sperm Whale ◽  
Heme Iron ◽  
Iron Protein ◽  
Protein Active Sites ◽  
Ligand Free ◽  
Ph Range ◽  
Protein Models ◽  
Exogenous Ligand

The composition of ferric exogenous ligand-free His93Gly sperm whale myoglobin (H93G Mb) at neutral pH has been determined by examination of the spectral properties of the protein over the pH range from 3.0 to 10.5. An apparent pKa value of ~6.6 has been observed for the conversion of a postulated six-coordinate bis-water-bound coordination structure at pH 5.0 to a five-coordinate hydroxide-bound form at pH 10.5. Starting from the exogenous ligand-free ferric H93G protein, ferric mono- and bis-thioether (tetrahydrothiophene, THT)-ligated adducts have been prepared and characterized by UV-visible (UV-vis) absorption and magnetic circular dichroism (MCD) spectroscopy. The mon-THT ferric H93G Mb species has hydroxide as the sixth ligand. The bis-THT derivative is a model for the low-spin ferric heme binding site of native bis-Met-ligated bacterioferritin or streptococcal heme-associated protein (Shp). A novel THT-bound ferryl H93G Mb moiety has been partially formed. The high-spin five-coordinate ferric H93G(selenolate) Mb complex has been prepared using benzeneselenol and characterized by UV-vis and MCD spectroscopy as a model for Se-Cys-ligated ferric cytochrome P450. The results described herein further demonstrate the versatility of the H93G cavity mutant for modeling the coordination structures of novel heme iron protein active sites.

Bindings of NO, CO, and O2 to multifunctional globin type dehaloperoxidase follow the ‘sliding scale rule’

2017 ◽  
Vol 474 (20) ◽  
pp. 3485-3498 ◽  
Author(s):  
Gang Wu ◽  
Jing Zhao ◽  
Stefan Franzen ◽  
Ah-Lim Tsai
Keyword(s):  
Oxygen Carrier ◽  
Sperm Whale ◽  
Dissociation Rate ◽  
Heme Iron ◽  
Binding Affinities ◽  
Ligand Selectivity ◽  
Sliding Scale ◽  
One Step ◽  
Sensor Proteins ◽  
Kinetics Of

Dehaloperoxidase–hemoglobin (DHP), a multifunctional globin protein, not only functions as an oxygen carrier as typical globins such as myoglobin and hemoglobin, but also as a peroxidase, a mono- and dioxygenase, peroxygenase, and an oxidase. Kinetics of DHP binding to NO, CO, and O2 were characterized for wild-type DHP A and B and the H55D and H55V DHP A mutants using stopped-flow methods. All three gaseous ligands bind to DHP significantly more weakly than sperm whale myoglobin (SWMb). Both CO and NO bind to DHP in a one-step process to form a stable six-coordinate complex. Multiple-step NO binding is not observed in DHP, which is similar to observations in SWMb, but in contrast with many heme sensor proteins. The weak affinity of DHP for O2 is mainly due to a fast O2 dissociation rate, in accordance with a longer εN–Fe distance between the heme iron and distal histidine in DHP than that in Mb, and an open-distal pocket that permits ligand escape. Binding affinities in DHP show the same 3–4 orders separation between the pairs NO/CO and CO/O2, consistent with the ‘sliding scale rule’ hypothesis. Strong gaseous ligand discrimination by DHP is very different from that observed in typical peroxidases, which show poor gaseous ligand selectivity, correlating with a neutral proximal imidazole ligand rather than an imidazolate. The present study provides useful insights into the rationale for DHP to function both as mono-oxygenase and oxidase, and is the first example of a globin peroxidase shown to follow the ‘sliding scale rule’ hypothesis in gaseous ligand discrimination.

scholarly journals Influence of the heme distal pocket on nitrite binding orientation and reactivity in Sperm Whale myoglobin

2021 ◽  
Vol 478 (4) ◽  
pp. 927-942
Author(s):  
Wilford Tse ◽  
Nathan Whitmore ◽  
Myles R. Cheesman ◽  
Nicholas J. Watmough
Keyword(s):  
Thermal Equilibrium ◽  
Magnetic Circular Dichroism ◽  
Sperm Whale ◽  
Heme Iron ◽  
Nitrite Reduction ◽  
Wild Type ◽  
Nitrite Ion ◽  
Alkaline Transition ◽  
Ferric Heme ◽  
Sperm Whale Myoglobin

Nitrite binding to recombinant wild-type Sperm Whale myoglobin (SWMb) was studied using a combination of spectroscopic methods including room-temperature magnetic circular dichroism. These revealed that the reactive species is free nitrous acid and the product of the reaction contains a nitrite ion bound to the ferric heme iron in the nitrito- (O-bound) orientation. This exists in a thermal equilibrium with a low-spin ground state and a high-spin excited state and is spectroscopically distinct from the purely low-spin nitro- (N-bound) species observed in the H64V SWMb variant. Substitution of the proximal heme ligand, histidine-93, with lysine yields a novel form of myoglobin (H93K) with enhanced reactivity towards nitrite. The nitrito-mode of binding to the ferric heme iron is retained in the H93K variant again as a thermal equilibrium of spin-states. This proximal substitution influences the heme distal pocket causing the pKa of the alkaline transition to be lowered relative to wild-type SWMb. This change in the environment of the distal pocket coupled with nitrito-binding is the most likely explanation for the 8-fold increase in the rate of nitrite reduction by H93K relative to WT SWMb.

Computational methods for calculation of protein-ligand binding affinities in structure-based drug design

2020 ◽  
Vol 0 (0) ◽  
Author(s):  
Zbigniew Dutkiewicz
Keyword(s):  
Drug Design ◽  
Ligand Binding ◽  
Binding Free Energy ◽  
Development Project ◽  
Quantum Mechanical ◽  
Prediction Methods ◽  
Binding Affinities ◽  
Structure Based Drug Design ◽  
Quantum Mechanical Description ◽  
Optimization Stage

AbstractDrug design is an expensive and time-consuming process. Any method that allows reducing the time the costs of the drug development project can have great practical value for the pharmaceutical industry. In structure-based drug design, affinity prediction methods are of great importance. The majority of methods used to predict binding free energy in protein-ligand complexes use molecular mechanics methods. However, many limitations of these methods in describing interactions exist. An attempt to go beyond these limits is the application of quantum-mechanical description for all or only part of the analyzed system. However, the extensive use of quantum mechanical (QM) approaches in drug discovery is still a demanding challenge. This chapter briefly reviews selected methods used to calculate protein-ligand binding affinity applied in virtual screening (VS), rescoring of docked poses, and lead optimization stage, including QM methods based on molecular simulations.

scholarly journals SMPLIP-Score: predicting ligand binding affinity from simple and interpretable on-the-fly interaction fingerprint pattern descriptors

2021 ◽  
Vol 13 (1) ◽  
Author(s):  
Surendra Kumar ◽  
Mi-hyun Kim
Keyword(s):  
Ligand Binding ◽  
Binding Affinity ◽  
Scoring Functions ◽  
Binding Affinities ◽  
Ligand Interaction ◽  
Fingerprint Pattern ◽  
Comparable Performance ◽  
Direct Interpretation ◽  
Benchmark Datasets ◽  
Complex Features

AbstractIn drug discovery, rapid and accurate prediction of protein–ligand binding affinities is a pivotal task for lead optimization with acceptable on-target potency as well as pharmacological efficacy. Furthermore, researchers hope for a high correlation between docking score and pose with key interactive residues, although scoring functions as free energy surrogates of protein–ligand complexes have failed to provide collinearity. Recently, various machine learning or deep learning methods have been proposed to overcome the drawbacks of scoring functions. Despite being highly accurate, their featurization process is complex and the meaning of the embedded features cannot directly be interpreted by human recognition without an additional feature analysis. Here, we propose SMPLIP-Score (Substructural Molecular and Protein–Ligand Interaction Pattern Score), a direct interpretable predictor of absolute binding affinity. Our simple featurization embeds the interaction fingerprint pattern on the ligand-binding site environment and molecular fragments of ligands into an input vectorized matrix for learning layers (random forest or deep neural network). Despite their less complex features than other state-of-the-art models, SMPLIP-Score achieved comparable performance, a Pearson’s correlation coefficient up to 0.80, and a root mean square error up to 1.18 in pK units with several benchmark datasets (PDBbind v.2015, Astex Diverse Set, CSAR NRC HiQ, FEP, PDBbind NMR, and CASF-2016). For this model, generality, predictive power, ranking power, and robustness were examined using direct interpretation of feature matrices for specific targets.

scholarly journals Novel Method for Quantifying AhR-Ligand Binding Affinities Using Microscale Thermophoresis

Biosensors ◽  
2021 ◽  
Vol 11 (3) ◽  
pp. 60
Author(s):  
Anne Stinn ◽  
Jens Furkert ◽  
Stefan H. E. Kaufmann ◽  
Pedro Moura-Alves ◽  
Michael Kolbe
Keyword(s):  
Ligand Binding ◽  
Environmental Pollutants ◽  
Screening Strategy ◽  
Pharmacological Intervention ◽  
Binding Affinities ◽  
Free System ◽  
Microscale Thermophoresis ◽  
Aryl Hydrocarbon ◽  
Promising Target ◽  
A Cell

The aryl hydrocarbon receptor (AhR) is a highly conserved cellular sensor of a variety of environmental pollutants and dietary-, cell- and microbiota-derived metabolites with important roles in fundamental biological processes. Deregulation of the AhR pathway is implicated in several diseases, including autoimmune diseases and cancer, rendering AhR a promising target for drug development and host-directed therapy. The pharmacological intervention of AhR processes requires detailed information about the ligand binding properties to allow specific targeting of a particular signaling process without affecting the remaining. Here, we present a novel microscale thermophoresis-based approach to monitoring the binding of purified recombinant human AhR to its natural ligands in a cell-free system. This approach facilitates a precise identification and characterization of unknown AhR ligands and represents a screening strategy for the discovery of potential selective AhR modulators.

scholarly journals Crystal Structure, Exogenous Ligand Binding, and Redox Properties of an Engineered Diiron Active Site in a Bacterial Hemerythrin

2013 ◽  
Vol 52 (22) ◽  
pp. 13014-13020 ◽  
Author(s):  
Yasunori Okamoto ◽  
Akira Onoda ◽  
Hiroshi Sugimoto ◽  
Yu Takano ◽  
Shun Hirota ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Ligand Binding ◽  
Active Site ◽  
Redox Properties ◽  
Exogenous Ligand

scholarly journals The role of Val68(E11) in ligand binding to sperm whale myoglobin. Site-directed mutagenesis of a synthetic gene.

1990 ◽  
Vol 265 (20) ◽  
pp. 11788-11795
Author(s):  
K D Egeberg ◽  
B A Springer ◽  
S G Sligar ◽  
T E Carver ◽  
R J Rohlfs ◽  
...  
Keyword(s):  
Ligand Binding ◽  
Sperm Whale ◽  
Site Directed Mutagenesis ◽  
Synthetic Gene ◽  
Directed Mutagenesis ◽  
Sperm Whale Myoglobin
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