Halogen Atoms in the Protein–Ligand System. Structural and Thermodynamic Studies of the Binding of Bromobenzotriazoles by the Catalytic Subunit of Human Protein Kinase CK2

Abstract

Binding of a family of brominated benzotriazoles to the catalytic subunit of human protein kinase CK2 (hCK2α) was used as a model system to assess the contribution of halogen bonding to protein–ligand interaction. CK2 is a constitutively active pleiotropic serine/threonine protein kinase that belongs to the CMGC group of eukaryotic protein kinases (EPKs). Due to the addiction of some cancer cells, CK2 is an attractive and well-characterized drug target. Halogenated benzotriazoles act as ATP-competitive inhibitors with unexpectedly good selectivity for CK2 over other EPKs. We have characterized the interaction of bromobenzotriazoles with hCK2α by X-ray crystallography, low-volume differential scanning fluorimetry, and isothermal titration calorimetry. Properties of free ligands in solution were additionally characterized by volumetric and RT-HPLC measurements. Thermodynamic data indicate that the affinity increases with bromo substitution, with greater contributions from 5- and 6-substituents than 4- and 7-substituents. Except for 4,7-disubstituted compounds, the bromobenzotriazoles adopt a canonical pose with the triazole close to lysine 68, which precludes halogen bonding. More highly substituted benzotriazoles adopt many additional noncanonical poses, presumably driven by a large hydrophobic contribution to binding. Some noncanonical ligand orientations allow the formation of halogen bonds with the hinge region. Consistent with a predominantly hydrophobic interaction, the isobaric heat capacity decreases upon ligand binding, the more so the higher the substitution.

Insight into the Binding and Hydrolytic Preferences of hNudt16 Based on Nucleotide Diphosphate Substrates

“…During the measurement, the samples were heated in a temperature range of 20-95 • C at a rate of 1 • C/min, and the measurements were performed with an excitation power of 30%. All of the collected data measured with using the above two DSF methods were analyzed according to the two-state transition numerical model described previously [51], using the Marquardt algorithm implemented in the Origin 2019 package (OriginLab, Northampton, MA, USA). We fitted the melting temperatures (T m ) globally for at least three independent experiments (but separately for different DSF methods).…”

Synthesis of Novel Halogenated Heterocycles Based on o-Phenylenediamine and Their Interactions with the Catalytic Subunit of Protein Kinase CK2

“…The majority of halogenated ligands target the ATP-binding site of a protein kinase; however, their poses differ significantly [ 19 , 20 , 21 , 22 , 23 ]. Recent studies have indicated that the binding mode of halogenated benzotriazoles and benzimidazoles is generally driven by a competition between a hydrogen bond or salt bridge and halogen bonding [ 20 ], while ligand hydrophobicity contributes significantly to the binding affinity [ 23 , 24 ]. Systematic structural studies on complexes of halogenated ligands pointed the preferred geometry [ 25 , 26 , 27 ] and topology [ 21 , 28 ] of a halogen bond in protein-ligand systems, however, the thermodynamic contribution of a halogen bond to free energy of ligand binding remains debatable, being estimated in the range of 0.2 [ 29 ] to 5 kcal/mol [ 30 ].…”

“…Kinase CK2 is also involved in DNA damage response and DNA repair pathways, which may additionally contribute to the regulation of cancer cell survival [8]. Despite the fact that the exact function of hCK2 in oncogenesis remains unknown and also its contribution The majority of halogenated ligands target the ATP-binding site of a protein kinase; however, their poses differ significantly [19][20][21][22][23]. Recent studies have indicated that the binding mode of halogenated benzotriazoles and benzimidazoles is generally driven by a competition between a hydrogen bond or salt bridge and halogen bonding [20], while ligand hydrophobicity contributes significantly to the binding affinity [23,24].…”

“…Despite the fact that the exact function of hCK2 in oncogenesis remains unknown and also its contribution The majority of halogenated ligands target the ATP-binding site of a protein kinase; however, their poses differ significantly [19][20][21][22][23]. Recent studies have indicated that the binding mode of halogenated benzotriazoles and benzimidazoles is generally driven by a competition between a hydrogen bond or salt bridge and halogen bonding [20], while ligand hydrophobicity contributes significantly to the binding affinity [23,24]. Systematic The majority of halogenated ligands target the ATP-binding site of a protein kinase; however, their poses differ significantly [19][20][21][22][23].…”

“…Recent studies have indicated that the binding mode of halogenated benzotriazoles and benzimidazoles is generally driven by a competition between a hydrogen bond or salt bridge and halogen bonding [20], while ligand hydrophobicity contributes significantly to the binding affinity [23,24]. Systematic The majority of halogenated ligands target the ATP-binding site of a protein kinase; however, their poses differ significantly [19][20][21][22][23]. Recent studies have indicated that the binding mode of halogenated benzotriazoles and benzimidazoles is generally driven by a competition between a hydrogen bond or salt bridge and halogen bonding [20], while ligand hydrophobicity contributes significantly to the binding affinity [23,24].…”