Location Is Everything: Influence of His-Tag Fusion Site on Properties of Adenylosuccinate Synthetase from .

The requirement for fast and dependable protein purification methods is constant, either for functional studies of natural proteins or for the production of biotechnological protein products. The original procedure has to be formulated for each individual protein, and this demanding task was significantly simplified by the introduction of affinity tags. adenylosuccinate synthetase (AdSS) is present in solution in a dynamic equilibrium of monomers and biologically active homodimers.

Effect of histidine protonation state on ligand binding at the ATP-binding site of human protein kinase CK2.

Histidine residues contribute to numerous molecular interactions, owing to their structure with the ionizable aromatic side chain with pK close to the physiological pH. Herein, we studied how the two histidine residues, His115 and His160 of the catalytic subunit of human protein kinase CK2, affect the binding of the halogenated heterocyclic ligands at the ATP-binding site. Thermodynamic studies on the interaction between five variants of hCK2α (WT protein and four histidine mutants) and three ionizable bromo-benzotriazoles and their conditionally non-ionizable benzimidazole counterparts were performed with nanoDSF, MST, and ITC.

The pursuit of new alternative ways to eradicate Helicobacter pylori continues: Detailed characterization of interactions in the adenylosuccinate synthetase active site.

Purine nucleotide synthesis is realised only through the salvage pathway in pathogenic bacterium Helicobacter pylori. Therefore, the enzymes of this pathway, among them also the adenylosuccinate synthetase (AdSS), present potential new drug targets. This paper describes characterization of His-tagged AdSS from H.

Competition between electrostatic interactions and halogen bonding in the protein-ligand system: structural and thermodynamic studies of 5,6-dibromobenzotriazole-hCK2α complexes.

CK2 is a member of the CMGC group of eukaryotic protein kinases and a cancer drug target. It can be efficiently inhibited by halogenated benzotriazoles and benzimidazoles. Depending on the scaffold, substitution pattern, and pH, these compounds are either neutral or anionic.

Improvement of native structure-based peptides as efficient inhibitors of protein-protein interactions of SARS-CoV-2 spike protein and human ACE2.

New pathogens responsible for novel human disease outbreaks in the last two decades are mainly the respiratory system viruses. Not different was the last pandemic episode, caused by infection of a severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). One of the extensively explored targets, in the recent scientific literature, as a possible way for rapid development of COVID-19 specific drug(s) is the interaction between the receptor-binding domain of the virus' spike (S) glycoprotein and human receptor angiotensin-converting enzyme 2 (ACE2).

5,6-diiodo-1H-benzotriazole: new TBBt analogue that minutely affects mitochondrial activity.

4,5,6,7-Tetrabromo-1H-benzotriazole is widely used as the reference ATP-competitive inhibitor of protein kinase CK2. Herein, we study its new analogs: 5,6-diiodo- and 5,6-diiodo-4,7-dibromo-1H-benzotriazole. We used biophysical (MST, ITC) and biochemical (enzymatic assay) methods to describe the interactions of halogenated benzotriazoles with the catalytic subunit of human protein kinase CK2 (hCK2α).

Calmodulin as Ca-Dependent Interactor of FTO Dioxygenase.

FTO is an -methyladenosine demethylase removing methyl groups from nucleic acids. Several studies indicate the creation of FTO complexes with other proteins. Here, we looked for regulatory proteins recognizing parts of the FTO dioxygenase region.

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

Protein kinase CK2 is a highly pleiotropic protein kinase capable of phosphorylating hundreds of protein substrates. It is involved in numerous cellular functions, including cell viability, apoptosis, cell proliferation and survival, angiogenesis, or ER-stress response. As CK2 activity is found perturbed in many pathological states, including cancers, it becomes an attractive target for the pharma.

Single tryptophan Y160W mutant of homooligomeric E. coli purine nucleoside phosphorylase implies that dimers forming the hexamer are functionally not equivalent.

E. coli purine nucleoside phosphorylase is a homohexamer, which structure, in the apo form, can be described as a trimer of dimers. Earlier studies suggested that ligand binding and kinetic properties are well described by two binding constants and two sets of kinetic constants.

Effect of Posttranslational Modifications on the Structure and Activity of FTO Demethylase.

The FTO protein is involved in a wide range of physiological processes, including adipogenesis and osteogenesis. This two-domain protein belongs to the AlkB family of 2-oxoglutarate (2-OG)- and Fe(II)-dependent dioxygenases, displaying -methyladenosine (-meA) demethylase activity. The aim of the study was to characterize the relationships between the structure and activity of FTO.

Native Structure-Based Peptides as Potential Protein-Protein Interaction Inhibitors of SARS-CoV-2 Spike Protein and Human ACE2 Receptor.

Severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2) is a positive-strand RNA virus that causes severe respiratory syndrome in humans, which is now referred to as coronavirus disease 2019 (COVID-19). Since December 2019, the new pathogen has rapidly spread globally, with over 65 million cases reported to the beginning of December 2020, including over 1.5 million deaths.

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.

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.

Diketopiperazine-Based, Flexible Tadalafil Analogues: Synthesis, Crystal Structures and Biological Activity Profile.

Phosphodiesterase 5 (PDE5) is one of the most extensively studied phosphodiesterases that is highly specific for cyclic-GMP hydrolysis. PDE5 became a target for drug development based on its efficacy for treatment of erectile dysfunction. In the present study, we synthesized four novel analogues of the phosphodiesterase type 5 (PDE5) inhibitor-tadalafil, which differs in (i) ligand flexibility (rigid structure of tadalafil vs.

The halogenation of natural flavonoids, baicalein and chrysin, enhances their affinity to human protein kinase CK2.

A series of halogenated derivatives of natural flavonoids: baicalein and chrysin were designed and investigated as possible ligands for the catalytic subunit of tumor-associated human kinase CK2. Thermal shift assay method, in silico modeling, and high-performance liquid chromatography-derived hydrophobicity together with IC values determined in biochemical assay were used to explain the ligand affinity to the catalytic subunit of human protein kinase CK2. Obtained results revealed that substitution of baicalein and chrysin with halogen atom increases their binding affinity to hCK2α, and for 8-chlorochrysin the observed effect is even stronger than for the reference CK2 inhibitor-4,5,6,7-tetrabromo-1H-benzotriazole.

Rational drug-design approach supported with thermodynamic studies - a peptide leader for the efficient bi-substrate inhibitor of protein kinase CK2.

Numerous inhibitors of protein kinases act on the basis of competition, targeting the ATP binding site. In this work, we present a procedure of rational design of a bi-substrate inhibitor, complemented with biophysical assays. The inhibitors of this type are commonly engineered by combining ligands carrying an ATP-like part with a peptide or peptide-mimicking fragment that determines specificity.