DypB peroxidase for aflatoxin removal: New insights into the toxin degradation process.

Aflatoxin B (AFB) is one of the most potent carcinogens and a widespread food and feed contaminant. As for other toxins, many efforts are devoted to find efficient and environmentally-friendly methods to degrade AFB, such as enzymatic treatments, thus improving the safety of food and feed products. In this regard, the dye decolorizing peroxidase of type B (DypB) can efficiently degrade AFB.

Antitarget, Anti-SARS-CoV-2 Leads, Drugs, and the Drug Discovery-Genetics Alliance Perspective.

The most advanced antiviral molecules addressing major SARS-CoV-2 targets (Main protease, Spike protein, and RNA polymerase), compared with proteins of other human pathogenic coronaviruses, may have a short-lasting clinical efficacy. Accumulating knowledge on the mechanisms underlying the target structural basis, its mutational progression, and the related biological significance to virus replication allows envisaging the development of better-targeted therapies in the context of COVID-19 epidemic and future coronavirus outbreaks. The identification of evolutionary patterns based solely on sequence information analysis for those targets can provide meaningful insights into the molecular basis of host-pathogen interactions and adaptation, leading to drug resistance phenomena.

Structural Basis of Parasitic HSP90 ATPase Inhibition by Small Molecules.

Protozoan parasites are responsible for several harmful and widespread human diseases that cause high morbidity and mortality. Currently available treatments have serious limitations due to poor efficiency, strong adverse effects, and high cost. Hence, the identification of new targets and the development of specific drug therapies against parasitic diseases are urgent needs.

Structural Characterization of Human Heat Shock Protein 90 N-Terminal Domain and Its Variants K112R and K112A in Complex with a Potent 1,2,3-Triazole-Based Inhibitor.

Heat shock protein 90 (Hsp90) is a ubiquitous molecular chaperone that stabilizes client proteins in a folded and functional state. It is composed of two identical and symmetrical subunits and each monomer consists of three domains, the N-terminal (NTD), the middle (MD), and the C-terminal domain (CTD). Since the chaperone activity requires ATP hydrolysis, molecules able to occupy the ATP-binding pocket in the NTD act as Hsp90 inhibitors, leading to client protein degradation and cell death.

A comprehensive approach to X-ray crystallography for drug discovery at a synchrotron facility - The example of Diamond Light Source.

A detailed understanding of the interactions between drugs and their targets is crucial to develop the best possible therapeutic agents. Structure-based drug design relies on the availability of high-resolution structures obtained primarily through X-ray crystallography. Collecting and analysing quickly a large quantity of structural data is crucial to accelerate drug discovery pipelines.

Fam20C-mediated phosphorylation of osteopontin is critical for its secretion but dispensable for its action as a cytokine in the activation of hepatic stellate cells in liver fibrogenesis.

Osteopontin (OPN) is a phosphoglycoprotein secreted into the extracellular matrix upon liver injury, acting as a cytokine stimulates the deposition of fibrillary collagen in liver fibrogenesis. In livers of mice subjected to bile duct ligation (BDL) and in cultured activated hepatic stellate cells (HSCs), we show that OPN, besides being overexpressed, is substantially phosphorylated by family with sequence similarity 20, member C (Fam20C), formerly known as Golgi casein kinase (G-CK), which is exclusively resident in the Golgi apparatus. In both experimental models, Fam20C becomes overactive when associated with a 500-kDa multiprotein complex, as compared with the negligible activity in livers of sham-operated rats and in quiescent HSCs.

Calcium-Induced Protein Folding in Calumenin and Calmodulin.

Binding of calcium - and small molecules in general - often induce conformational changes in large molecules and complexes. The degree and type of change varies, but the resulting shift in specific affinities ultimately induces a physiological response. It is therefore important for our understanding of responses at the cellular level to define coupled changes at the molecular level.

VMXi: a fully automated, fully remote, high-flux in situ macromolecular crystallography beamline.

VMXi is a new high-flux microfocus macromolecular crystallography beamline at Diamond Light Source. The beamline, dedicated to fully automated and fully remote data collection of macromolecular crystals in situ, allows rapid screening of hundreds of crystallization plates from multiple user groups. Its main purpose is to give fast feedback at the complex stages of crystallization and crystal optimization, but it also enables data collection of small and delicate samples that are particularly difficult to harvest using conventional cryo-methods, crystals grown in the lipidic cubic phase, and allows for multi-crystal data collections in drug discovery programs.

A generic protocol for protein crystal dehydration using the HC1b humidity controller.

Dehydration may change the crystal lattice and affect the mosaicity, resolution and quality of X-ray diffraction data. A dehydrating environment can be generated around a crystal in several ways with various degrees of precision and complexity. This study uses a high-precision crystal humidifier/dehumidifier to provide an airstream of known relative humidity in which the crystals are mounted: a precise yet hassle-free approach to altering crystal hydration.

Ca-Dependent Folding of Human Calumenin.

Human calumenin (hCALU) is a six EF-hand protein belonging to the CREC family. As other members of the family, it is localized in the secretory pathway and regulates the activity of SERCA2a and of the ryanodine receptor in the endoplasmic reticulum (ER). We have studied the effects of Ca2+ binding to the protein and found it to attain a more compact structure upon ion binding.

Application of in situ diffraction in high-throughput structure determination platforms.

Macromolecular crystallography (MX) is the most powerful technique available to structural biologists to visualize in atomic detail the macromolecular machinery of the cell. Since the emergence of structural genomics initiatives, significant advances have been made in all key steps of the structure determination process. In particular, third-generation synchrotron sources and the application of highly automated approaches to data acquisition and analysis at these facilities have been the major factors in the rate of increase of macromolecular structures determined annually.

An evaluation of adhesive sample holders for advanced crystallographic experiments.

The hydration state of macromolecular crystals often affects their overall order and, ultimately, the quality of the X-ray diffraction pattern that they produce. Post-crystallization techniques that alter the solvent content of a crystal may induce rearrangement within the three-dimensional array making up the crystal, possibly resulting in more ordered packing. The hydration state of a crystal can be manipulated by exposing it to a stream of air at controlled relative humidity in which the crystal can equilibrate.

Lyn-mediated procaspase 8 dimerization blocks apoptotic signaling in B-cell chronic lymphocytic leukemia.

Lyn, a member of the group of tyrosine kinases named the Src family kinases (SFKs), is overexpressed, associated with an aberrant multiprotein complex and constitutively active in B-cell chronic lymphocytic leukemia (B-CLL) cells, resulting in a high level of tyrosine phosphorylation and contributing to their resistance to apoptosis. By using biochemical and bioinformatics tools, we identified procaspase-8 (procasp8), the caspase-8 zymogen, as a cytosolic target for Lyn in B-CLL cells, the phosphorylation of which at Tyr380 promotes the formation of an inactive procasp8 homodimer. This complex remains segregated in the cytosol and appears to be crucial in mediating the antiapoptotic function of Lyn in this disease.

Comparative transcriptomics reveals RhoE as a novel regulator of actin dynamics in bone-resorbing osteoclasts.

The function of osteoclasts (OCs), multinucleated giant cells (MGCs) of the monocytic lineage, is bone resorption. To resorb bone, OCs form podosomes. These are actin-rich adhesive structures that pattern into rings that drive OC migration and into "sealing-zones" (SZs) that confine the resorption lacuna.

DAP12 overexpression induces osteopenia and impaired early hematopoiesis.

ITAM-bearing transmembrane signaling adaptors such as DAP12 and FcRγ are important players in bone homeostasis, but their precise role and functions are still unknown. It has been shown that osteoclast differentiation results from the integration of the RANK and of the DAP12 and FcRγ signaling pathways. DAP12-deficient mice suffer from a mild osteopetrosis and culture of their bone marrow cells in the presence of M-CSF and RANKL, fails to give rise to multinucleated osteoclasts.

Chemoresistance of human monocyte-derived dendritic cells is regulated by IL-17A.

Dendritic cells initiate adaptive immune responses, leading either to control cancer by effector T cells or to exacerbate cancer by regulatory T cells that inhibit IFN-γ-mediated Th1-type response. Dendritic cells can also induce Th17-type immunity, mediated by IL-17A. However, the controversial role of this cytokine in cancer requires further investigations.

Inhibition of protein kinase CK2 by flavonoids and tyrphostins. A structural insight.

Sixteen flavonoids and related compounds have been tested for their ability to inhibit three acidophilic Ser/Thr protein kinases: the Golgi apparatus casein kinase (G-CK) recently identified with protein FAM20C, protein kinase CK1, and protein kinase CK2. While G-CK is entirely insensitive to all compounds up to 40 μM concentration, consistent with the view that it is not a member of the kinome, and CK1 is variably inhibited in an isoform-dependent manner by fisetin and luteolin, and to a lesser extent by myricetin and quercetin, CK2 is susceptible to drastic inhibition by many flavonoids, displaying with six of them IC(50) values < 1 μM. A common denominator of these compounds (myricetin, quercetin, fisetin, kaempferol, luteolin, and apigenin) is a flavone scaffold with at least two hydroxyl groups at positions 7 and 4'.

Hit to lead evaluation of 1,2,3-triazolo[4,5-b]pyridines as PIM kinase inhibitors.

PIM kinases have become targets of interest due to their association with biochemical mechanisms affecting survival, proliferation and cytokine production. 1,2,3-Triazolo[4,5-b]pyridines were identified as PIM inhibitors applying a scaffold hopping approach. Initial exploration around this scaffold and X-ray crystallographic data are hereby described.

Structural features underlying the selectivity of the kinase inhibitors NBC and dNBC: role of a nitro group that discriminates between CK2 and DYRK1A.

8-hydroxy-4-methyl-9-nitrobenzo(g)chromen-2-one (NBC) has been found to be a fairly potent ATP site-directed inhibitor of protein kinase CK2 (Ki = 0.22 μM). Here, we show that NBC also inhibits PIM kinases, especially PIM1 and PIM3, the latter as potently as CK2.

The centrosome: a target for cancer therapy.

The centrosome plays an essential role in cell cycle progression and cell polarity, organizing the microtubule network in interphase and mitosis. During cell division, the centrosome undergoes a series of structural and functional transitions and forms the two poles of the bipolar mitotic spindle. It is the microtubule cytoskeleton that is reorganized to form the two poles, ensuring accurate separation of the two daughter cells.

The pleiotropic protein kinase CK2 phosphorylates HTLV-1 Tax protein in vitro, targeting its PDZ-binding motif.

The HTLV-1 transactivator Tax is an oncoprotein capable of deregulating the expression of many cellular genes and interfering with signalling pathways. Here we show that Tax-1 is phosphorylated in vitro by the pleiotropic human serine/threonine kinase CK2 at three residues, Ser-336, Ser-344 and Thr-351, close to and within its C-terminal PDZ-binding motif. We also show that the mutation of Thr-351 to aspartate abolishes Tax-1 binding to the scaffold protein hDlg, a tumour suppressor factor, while having no effect on transactivation.

Genome-wide expression analyses establish dendritic cells as a new osteoclast precursor able to generate bone-resorbing cells more efficiently than monocytes.

Dendritic cells (DCs), mononuclear cells that initiate immune responses, and osteoclasts (OCs), multinucleated bone-resorbing cells, are hematopoietic cells derived from monocytic precursor cells. Using in vitro generated dendritic cells, we previously showed that human and murine DCs could transdifferentiate into resorbing osteoclasts in the presence of macrophage colony-stimulating factor (M-CSF) and receptor activator of NF-kappaB ligand (RANKL). In this study we globally compared by transcriptomic profiling this new osteoclast differentiation pathway from DCs with the canonical differentiation pathway from monocytes.

Quinalizarin as a potent, selective and cell-permeable inhibitor of protein kinase CK2.

Emodin (1,3,8-trihydroxy-6-methyl-anthraquinone) is a moderately potent and poorly selective inhibitor of protein kinase CK2, one of the most pleiotropic serine/threonine protein kinases, implicated in neoplasia and in other global diseases. By virtual screening of the MMS (Molecular Modeling Section) database, we have now identified quinalizarin (1,2,5,8-tetrahydroxyanthraquinone) as an inhibitor of CK2 that is more potent and selective than emodin. CK2 inhibition by quinalizarin is competitive with respect to ATP, with a Ki value of approx.

[Osteoimmunology: an integrated vision of immune and bone systems].

Study of dysregulations of the immune system resulting in abnormal bone homeostasis has led to the emergence of the field of osteoimmunology. Among the interactions between the immune system and bone, several signaling molecules and pathways previously identified in immune cells have been shown to be key players in the osteoclast, the bone resorbing cell. Osteoclast differentiation requires two signaling pathways: the RANK/RANKL pathway and the signal initiated by the ITAM-harboring adaptor proteins DAP12 and FcRgamma.

A structural insight into CK2 inhibition.

The acidophilic Ser/Thr protein kinase CK2 displays some unique properties such as high pleiotropicity and constitutive activity. CK2 is involved in many fundamental aspects of the normal cell life, for instance it promotes cell survival and enhances the tumour phenotype under special circumstances. This makes CK2 an appealing target for the development of inhibitors with pharmacological potential.