31 research outputs found
Dissection of the dual function of the β-subunit of protein kinase CK2 (‘casein kinase-2’): a synthetic peptide reproducing the carboxyl-terminal domain mimicks the positive but not the negative effects of the whole protein
No full textThe dual function of the regulatory beta-subunit of protein kinase CK2 is highlighted by its ability to abolish calmodulin phosphorylation in contrast to its stimulatory effect on the phosphorylation of peptide substrates, Here we show that a synthetic peptide reproducing the C-terminal region of the beta-subunit (beta[170-215]) stimulates to a similar extent the phosphorylation of either the peptide substrate or calmodulin and also protects the catalytic alpha-subunit against thermal inactivation as efficiently as full-length beta-subunit. These data show that the positive and negative functions of the beta-subunit reside in physically separated domains and that the elements responsible for positive regulation are located in the C-terminal region
Synthesis of Triazenoazaindoles: a New Class of Triazenes with Antitumor Activity
No full textDespite improvements in the treatment and prevention of cancer, the number of new diagnoses continues to rise; this has fuelled substantial interest in the development of new and effective chemotherapeutic agents. Compounds of the triazene class, such as dacarbazine, have been used in the clinical management
of many cancer types including brain, leukemia, and melanoma. A new compound class bearing a triazenoazaindole
scaffold was synthesized with the aim of identifying new antiproliferative
agents. Compounds 5a–g and 6a–c were
screened against a panel of human tumor cell lines, and two of them, 5e and 5 f, showed cytotoxicity (GI50 range: 2.2–8.2 mm) in all cell lines. These two compounds even maintained their cytotoxicity in some multidrug-resistant cell lines. Flow cytometry analysis demonstrated their ability to induce cell death by apoptosis with involvement of lysosomes
2-Triazenoazaindoles: Á novel class of triazenes inducing transcriptional down-regulation of EGFR and HER-2 in human pancreatic cancer cells
No full textPancreatic cancer is a complex malignancy arising from the accumulation of genetic and epigenetic defects in the affected cells. Standard chemotherapy for patients with advanced disease shows only modest effects and is associated with considerable toxicity. Overexpression or aberrant activation of members of the epidermal growth factor receptor tyrosine kinase family, which includes EGFR and HER-2, occurs frequently and is associated with multiple drug resistance and decreased patient survival. In this study, we have investigated the therapeutic potential of AS104, a novel compound of the triazene class, with potential inhibitory effects on EGFR. We found that treatment of cells with AS104 causes significant reduction of cell growth and metabolic activity in four human pancreatic cancer cell lines. Furthermore, we show that the AS104-mediated induction of apoptotic cell death is associated with stimulation of autophagy in a dose-dependent manner. Treatment of cells with AS104 results in significant down-regulation of EGFR and HER-2 expression and activity and subsequent inhibition of downstream signaling proteins. Quantitative RT-PCR analysis and assays with proteasome inhibitors revealed that AS104 regulates the expression of EGFR and HER-2 at the transcriptional level. These findings provide for the first time experimental evidence for efficacy of AS104 in the simultaneous transcriptional repression of EGFR and HER-2 genes and suggest that AS104 may have therapeutic potential in the treatment of pancreatic cancers that express high levels of the aforementioned receptor tyrosine kinase
A-Raf kinase is a new interacting partner of protein kinase CK2 β subunit
No full textAbstractIn a search for protein kinase CK2 β subunit binding proteins using the two-hybrid system, more than 1000 positive clones were isolated. Beside clones for the α′ and β subunit of CK2, there were clones coding for a so far unknown protein, whose partial cDNA sequence was already deposited in the EMBL database under the accession numbers R08806 and Z17360, for the ribosomal protein L5 and for A-Raf kinase. All isolated clones except the one for CK2 β showed no interaction with the catalytic α subunit of CK2. A-Raf kinase is a new interesting partner of CK2 β. The isolated A-Raf clone represented amino acids 268–606, but also a full length A-Raf clone interacted with CK2 β. At the site of CK2 β, residue 175 and amino acids between residues 194 and 200 are likely to be involved in direct interaction.© 1997 Federation of European Biochemical Societies
p53 and the ribosomal protein L5 participate in high molecular mass complex formation with protein kinase CK2 in murine teratocarcinoma cell line F9 after serum stimulation and cisplatin treatment
No full textAbstractUsing the murine teratocarcinoma cell line F9 we investigated the influence of serum stimulation and cisplatin treatment on the p53, CK2, MDM2 levels. Both treatments led to an increase of p53, though with different kinetics; the other proteins investigated were not affected. We present direct evidence by immunoprecipitation for an association of protein kinase CK2 holoenzyme (α2β2), p53, and the ribosomal protein L5. The results suggest complexes between the CK2 holoenzyme and p53 but also p53/CKβ complexes. Furthermore we provide evidence for the existence of high molecular mass complexes of CK2 in vivo. This is the first evidence that, under physiological conditions, protein kinase CK2 does not exist solely as a heterotetramer, but predominantly in association with other proteins
Synthesis of a new class of pyrrolo[3,4-h]quinazolines with antimitotic activity
No full textA new series of pyrrolo[3,4-h]quinazolines was conveniently prepared with a broad substitution pattern.
A large number of derivatives was obtained and the cellular cytotoxicity was evaluated in vitro against 5
different human tumor cell lines with GI50 values reaching the low micromolar level (1.3e19.8 mM).
These compounds were able to induce cell death mainly by apoptosis through a mitochondrial dependent
pathway. Selected compounds showed antimitotic activity and a reduction of tubulin polymerization
in a concentration-dependent manner. Moreover, they showed anti-angiogenic properties since
reduced in vitro endothelial cell migration and disrupted HUVEC capillary-like tube network in Matrigel
Low density crystal packing of human protein kinase CK2 catalytic subunit in complex with resorufin or other ligands a tool to study the unique hinge region plasticity of the enzyme without packing bias
No full textA low-resolution structure of the catalytic subunit CK2α of human protein kinase CK2 (formerly known as casein kinase 2) in complex with the ATP-competitive inhibitor resorufin is presented. The structure supplements previous human CK2α structures in which the interdomain hinge/helix αD region adopts a closed conformation correlating to a canonically established catalytic spine as is typical for eukaryotic protein kinases. In the corresponding crystal packing the hinge/helix αD region is nearly unaffected by crystal contacts, so that largely unbiased conformational adaptions are possible. This is documented by published human CK2α structures with the same crystal packing but with an open hinge/helix αD region, one of which has been redetermined here with a higher symmetry. An overview of all published human CK2α crystal packings serves as the basis for a discussion of the factors that determine whether the open or the closed hinge/helix αD conformation is adopted. Lyotropic salts in crystallization support the closed conformation, in which the Phe121 side chain complements the hydrophobic catalytic spine ensemble. Consequently, genuine ligand effects on the hinge/helix αD conformation can be best studied under moderate salt conditions. Ligands that stabilize either the open or the closed conformation by hydrogen bonds are known, but a general rule is not yet apparent
Insights from soft X-rays: the chlorine and sulfur sub-structures of a CK2alpha/DRB complex
No full textThe diffraction pattern of a protein crystal is normally a product of the interference of electromagnetic waves scattered by electrons of the crystalline sample. The diffraction pattern undergoes systematic changes in case additionally X-ray absorption occurs, meaning if the wavelength of the primary X-ray beam is relatively close to the absorption edge of selected elements of the sample. The resulting effects are summarized as "anomalous dispersion" and can be always observed with "soft" X-rays (wavelength around 2 A) since they match the absorption edges of sulfur and chlorine. A particularly useful application of this phenomenon is the experimental detection of the sub-structures of the anomalous scatterers in protein crystals. We demonstrate this here with a crystal of a C-terminally truncated variant of human CK2alpha to which two molecules of the inhibitor 5,6-dichloro-1-beta-D-ribo-furanosyl-benzimidazole (DRB) are bound. The structure of this co-crystal has been solved recently. For this study we measured an additional diffraction data set at a wavelength of 2 A which showed strong anomalous dispersion effects. On the basis of these effects we detected all sulfur atoms of the protein, the two liganded DRB molecules and a total of 16 additional chloride ions some of them emerging at positions filled with water molecules in previous structure determinations. A number of chloride ions are bound to structural and functional important locations fitting to the constitutive activity and the acidophilic substrate specificity of the enzyme
A Casein-Kinase-2-Related Protein Kinase is Tightly Associated with the Large T Antigen of Simian Virus 40
No full textFirst inactive conformation of CK2alpha, the catalytic subunit of protein kinase CK2
No full textUdgivelsesdato: 2009-Mar-13The Ser/Thr kinase casein kinase 2 (CK2) is a heterotetrameric enzyme composed of two catalytic chains (CK2alpha, catalytic subunit of CK2) attached to a dimer of two noncatalytic subunits (CK2beta, noncatalytic subunit of CK2). CK2alpha belongs to the superfamily of eukaryotic protein kinases (EPKs). To function as regulatory key components, EPKs normally exist in inactive ground states and are activated only upon specific signals. Typically, this activation is accompanied by large conformational changes in helix alpha C and in the activation segment, leading to a characteristic arrangement of catalytic key elements. For CK2alpha, however, no strict physiological control of activity is known. Accordingly, CK2alpha was found so far exclusively in the characteristic conformation of active EPKs, which is, in this case, additionally stabilized by a unique intramolecular contact between the N-terminal segment on one side, and helix alpha C and the activation segment on the other side. We report here the structure of a C-terminally truncated variant of human CK2alpha in which the enzyme adopts a decidedly inactive conformation for the first time. In this CK2alpha structure, those regulatory key regions still are in their active positions. Yet the glycine-rich ATP-binding loop, which is normally part of the canonical anti-parallel beta-sheet, has collapsed into the ATP-binding site so that ATP is excluded from binding; specifically, the side chain of Arg47 occupies the ribose region of the ATP site and Tyr50, the space required by the triphospho moiety. We discuss some factors that may support or disfavor this inactive conformation, among them coordination of small molecules at a remote cavity at the CK2alpha/CK2beta interaction region and binding of a CK2beta dimer. The latter stabilizes the glycine-rich loop in the extended active conformation known from the majority of CK2alpha structures. Thus, the novel inactive conformation for the first time provides a structural basis for the stimulatory impact of CK2beta on CK2alpha
