1,720,996 research outputs found
Structural dissection of cyclin dependent kinases regulation and protein recognition properties.
No full textCyclin Dependent Kinases (CDKs) regulate the cell division cycle, apoptosis, transcription and differentiation in addition to functions in the nervous system. They are regulated by their cyclin partners and by a variety of additional protein effectors (inhibitors, kinases, phosphatases). Each CDK serves its function by means of specific protein recognition properties. These are also responsible for the differential regulation of CDKs/Cyclin couples involved in processes as different as cell cycle and transcription. The structural features determining general and specific properties for CDKs/Cyclin complexes are analyzed. They reside in an overall conserved architecture with divergent spots used by the complexes to present themselves to specific substrates or other protein effectors
Binding to DNA of the RNA-polymerase II C-terminal domain allows discrimination between Cdk7 and Cdk9 phosphorylation
Full text linkThe C-terminal domain (CTD) of RNA polymerase II regulates transcription through spatially and temporally coordinated events. Previous work had established that the CTD binds DNA but the significance of this interaction had not been determined. The present work shows that the CTD binds DNA in its unphosphorylated form, the form in which it is present in the pre-initiation complex. The CTD/DNA complex is recognized by and is phosphorylated by Cdk7 but not by Cdk9. Model-building studies indicate the structural mechanism underlying such specificity involves interaction of Cdk7 with DNA in the context of the CTD/DNA complex. The model has been tested by mutagenesis experiments. CTD dissociates from DNA following phosphorylation by Cdk7, allowing transcription initiation. The CTD then becomes accessible for further phosphorylation by Cdk9 that drives the transition to transcription elongation
Different orientations of low-molecular-weight fragments in the binding pocket of a BRD4 bromodomain
No full textBromodomains are involved in the regulation of chromatin architecture and transcription through the recognition of acetylated lysines in histones and other proteins. Many of them are considered to be relevant pharmacological targets for different pathologies. Three crystallographic structures of the N-terminal bromodomain of BRD4 in complex with low-molecular-weight fragments are presented. They show that similar molecules mimicking acetylated lysine bind the bromodomain with different orientations and exploit different interactions. It is also advised to avoid DMSO when searching for low-affinity fragments that interact with bromodomains since DMSO binds in the acetylated lysine-recognition pocket of BRD4
CAK—Cyclin-Dependent Activating Kinase: A Key Kinase in Cell Cycle Control and a Target for Drugs?
No full textThe Cyclin-dependent kinase (CDK) Activating Kinase (CAK) is responsible for the activating phosphorylation of CDK1, CDK2, CDK4 and CDK6 and regulation of the cell cycle. The kinase is composed of three subunits: CDK7, Cyclin H and MAT1 (ménage a trois). Together with six other subunits, CAK is also part of the general transcription factor TFIIH where it is involved in promoter clearance and progression of transcription from the preinitiation to the initiation stage. CAK is required for cell cycle progression, which suggests that CDK7 could be a target for cancer therapy. However its role in transcription and its ubiquitous presence raise sensible concerns about possible toxicity of its inhibitors. The recently determined structure of CDK7 allows the design of inhibitors with differential specificity for the different CDKs. We review the role of CAK in different biological processes and evaluate the biological evidence for CDK7 as a possible pharmacological target
Recognition of Cdk2 by Cdk7
No full textCdk7, a member of the cyclin dependent protein kinase family, regulates the activities of other Cdks through phosphorylation on their activation segment, and hence contributes to control of the eukaryotic cell cycle. Cdk7 is itself phosphorylated on the activation segment. Cdk7 phosphorylates Cdk1, Cdk2, Cdk4, and Cdk6, but only Cdk1 and Cdk2 can phosphorylate Cdk7 and none of them is able to auto-phosphorylate. The activation segments of the Cdks are very similar in sequence. Their specificity does not appear to be dictated by the sequences surrounding the phosphorylation sites but by structural determinants at remote sites. Through mutagenesis studies, we have identified regions in Cdk2 responsible for its interaction with Cdk7. A model has been built that explains the molecular basis for the specificity observed in Cdk recognition. The two kinases are arranged in a quasi-symmetric head-to-tail arrangement in which the N-terminal lobe from one kinase docks against the C-terminal lobe from the other kinase, and the activation segments are within reach of the opposite catalytic sites. Further experiments demonstrate that cyclin A hydrophobic pocket is not a recruitment site for Cdk7
Active Form of the Protein Kinase CK2 alpha(2)beta(2) holoenzyme Is a Strong Complex with Symmetric Architecture
No full textCK2 is a protein kinase essential for cell viability whose activity is altered in several cancers. Its mechanisms of regulation differ from those common to other eukaryotic protein kinases and are not entirely established yet. Here we present crystal structures of the monomeric form of the alpha(2)beta(2) holoenzyme that allow refining a formerly proposed structural model for activity regulation by oligomerization. Previous crystal structures of the CK2 holoenzyme show an asymmetric arrangement of the two alpha catalytic subunits around the obligate beta(2) regulatory subunits. Asymmetric alpha(2)beta(2) tetramers are organized in trimeric rings that correspond to inactive forms of the enzyme. The new crystal structures presented here reveal the symmetric architecture of the isolated active tetramers. The dimension and the nature of the alpha/beta interfaces configure the holoenzyme as a strong complex that does not spontaneously dissociate in solution, in accordance with the low dissociation constant (similar to 4 nM)
Structural Determinants of Protein Kinase CK2 Regulation by Autoinhibitory Polymerization
No full textCK2 is a Ser/Thr protein kinase essential for cell viability whose activity is anomalously high in several cancers. CK2 is a validated target for cancer therapy with one small molecule inhibitor in phase I clinical trials. This enzyme is not regulated by mechanisms common to other protein kinases, and how its activity is controlled is still unclear. We present a new crystal structure of the CK2 holoenzyme that supports an autoinhibitory mechanism of regulation whereby the β-subunit plays an essential role in the formation of inactive polymeric assemblies. The derived structural model of (down)regulation by aggregation contributes to the interpretation of biochemical and functional data and paves the way for new strategies in the modulation of CK2 activity and for the design of non-ATP-competitive inhibitors targeting the interaction between the α catalytic and the β regulatory subunits
Structural and functional determinants of protein kinase CK2 alpha: facts and open questions
No full textSer/Thr protein kinase CK2 is involved in several fundamental processes that regulate the cell life, such as cell cycle progression, gene expression, cell growth, and differentiation and embryogenesis. In various cancers, CK2 shows a markedly elevated activity that has been associated with conditions that favor the onset of the tumor phenotype. This prompts to numerous studies aimed at the identification of compounds that are able to inhibit the catalytic activity of this oncogenic kinase, in particular, of ATP-competitive inhibitors. The many available crystal structures indicate that this enzyme owns some regions of remarkable flexibility which were associated to important functional properties. Of particular relevance is the flexibility, unique among protein kinases, of the hinge region and the following helix alpha D. This study attempts to unveil the structural bases of this characteristic of CK2. We also analyze some controversial issues concerning the functional interpretation of structural data on maize and human CK2 and try to recognize what is reasonably established and what is still unclear about this enzyme. This analysis can be useful also to outline some principles at the basis of the development of effective ATP-competitive CK2 inhibitors
The STAS domain of mammalian SLC26A5 prestin harbours an anion-binding site
No full textPrestin is a unique ATP-and Ca2+-independent molecular motor with piezoelectric characteristics responsible for the electromotile properties ofmammalian cochlear outer hair cells, i.e.The capacity of these cells to modify their length in response to electric stimuli. This 'electromotility' is at the basis of the exceptional sensitivity and frequency selectivity distinctive of mammals. Prestin belongs to the SLC26 (solute carrier 26) family of anion transporters and needs anions to function properly, particularly Cl? . In the present study, using X-ray crystallography we reveal that the STAS (sulfate transporter and anti-sigma factor antagonist) domain of mammalian prestin, considered an 'incomplete' transporter, harbours an unanticipated anion-binding site. In parallel, we present the first crystal structure of a prestin STAS domain from a non-mammalian vertebrate prestin (chicken) that behaves as a 'full' transporter. Notably, in chicken STAS, the anion-binding site is lacking because of a local structural rearrangement, indicating that the presence of the STAS anion-binding site is exclusive to mammalian prestin. © 2016 Authors; published by Portland Press Limited
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