Relevance of the paper is due to the need for understanding of educational environment, conducive to the formation of a creative, holistic and harmonious personality. The study used historical and ...cultural methods, method of comparative and system analysis. In this regard, complementary relationship between humanities and natural sciences, contributing to the development of symbolic thinking, was revealed. Peculiarities of symbolic thinking and its role in training the creative professional were shown. Approaches to investigating dialogue mode in the educational environment, resulting in the formation and harmonious development of the creative personality, were identified.
The p90 ribosomal S6 kinases (RSKs) also known as MAPKAP‐Ks are serine/threonine protein kinases that are activated by ERK or PDK1 and act as downstream effectors of mitogen‐activated protein kinase ...(MAPK). RSK1, a member of the RSK family, contains two distinct kinase domains in a single polypeptide chain, the regulatory C‐terminal kinase domain (CTKD) and the catalytic N‐terminal kinase domain (NTKD). Autophosphorylation of the CTKD leads to activation of the NTKD that subsequently phosphorylates downstream substrates. Here we report the crystal structures of the unactivated RSK1 NTKD bound to different ligands at 2.0 Å resolution. The activation loop and helix αC, key regulatory elements of kinase function, are disordered. The DFG motif of the inactive RSK1 adopts an “active‐like” conformation. The β‐PO4 group in the AMP–PCP complex adopts a unique conformation that may contribute to inactivity of the enzyme. Structures of RSK1 ligand complexes offer insights into the design of novel anticancer agents and into the regulation of the catalytic activity of RSKs.
The receptor tyrosine kinase c-Met is implicated in oncogenesis and is the target for several small molecule and biologic agents in clinical trials for the treatment of cancer. Binding of the ...hepatocyte growth factor to the cell surface receptor of c-Met induces activation via autophosphorylation of the kinase domain. Here we describe the structural basis of c-Met activation upon autophosphorylation and the selective small molecule inhibiton of autophosphorylated c-Met. MK-2461 is a potent c-Met inhibitor that is selective for the phosphorylated state of the enzyme. Compound 1 is an MK-2461 analog with a 20-fold enthalpy-driven preference for the autophosphorylated over unphosphorylated c-Met kinase domain. The crystal structure of the unbound kinase domain phosphorylated at Tyr-1234 and Tyr-1235 shows that activation loop phosphorylation leads to the ejection and disorder of the activation loop and rearrangement of helix αC and the G loop to generate a viable active site. Helix αC adopts a orientation different from that seen in activation loop mutants. The crystal structure of the complex formed by the autophosphorylated c-Met kinase domain and compound 1 reveals a significant induced fit conformational change of the G loop and ordering of the activation loop, explaining the selectivity of compound 1 for the autophosphorylated state. The results highlight the role of structural plasticity within the kinase domain in imparting the specificity of ligand binding and provide the framework for structure-guided design of activated c-Met inhibitors.
The x-ray structure of the unactivated kinase domain of insulin-like growth factor-1 receptor (IGFRK-0P) is reported here at 2.7 A resolution. IGFRK-0P is composed of two lobes connected by a hinge ...region. The N-terminal lobe of the kinase is a twisted beta-sheet flanked by a single helix, and the C-terminal lobe comprises eight alpha-helices and four short beta-strands. The ATP binding pocket and the catalytic center reside at the interface of the two lobes. Despite the overall similarity to other receptor tyrosine kinases, three notable conformational modifications are observed: 1) this kinase adopts a more closed structure, with its two lobes rotated further toward each other; 2) the conformation of the proximal end of the activation loop (residues 1121-1129) is different; 3) the orientation of the nucleotide-binding loop is altered. Collectively, these alterations lead to a different ATP-binding pocket that might impact on inhibitor designs for IGFRK-0P. Two molecules of IGFRK-0P are seen in the asymmetric unit; they are associated as a dimer with their ATP binding clefts facing each other. The ordered N terminus of one monomer approaches the active site of the other, suggesting that the juxtamembrane region of one molecule could come into close proximity to the active site of the other.
An optimal control problem with state constraints based on a SEIR model to control the spreading of infectious diseases is consi- dered. The main purpose is apply novel theoretical results to ...successfully validate the numerical solution, computed via direct method. The problem has simple but yet interesting features that we explore in our analysis. Of particular interest is the fact that the state constraint is of order one and that the solution is normal.
The type 1 insulin-like growth factor receptor (IGF-1R) is often overexpressed on tumor cells and is believed to play an important role in anchorage-independent proliferation. Additionally, cell ...culture studies have indicated that IGF-1R confers increased resistance to apoptosis caused by radiation or chemotherapeutic agents. Thus, inhibitors of the intracellular kinase domain of this receptor may have utility for the clinical treatment of cancer. As part of an effort to develop clinically useful inhibitors of IGF-1R kinase, a novel class of pyrrole-5-carboxaldehyde compounds was investigated. The compounds exhibited selectivity against the closely related insulin receptor kinase intrinsically and in cell-based assays. The inhibitors formed a reversible, covalent adduct at the kinase active site, and treatment of such adducts with sodium borohydride irreversibly inactivated the enzyme. Analysis of a tryptic digest of a covalently modified IGF-1R kinase fragment revealed that the active site Lys1003 had been reductively alkylated with the aldehyde inhibitor. Reductive alkylation of the insulin receptor kinase with one of these inhibitors led to a similarly inactivated enzyme which was examined by X-ray crystallography. The crystal structure confirmed the modification of the active site lysine side chain and revealed details of the key interactions between the inhibitor and enzyme.
The crystal structure of the wild‐type unactivated kinase domain (IGFRK‐0P) of insulin‐like growth factor‐1 receptor has been reported previously at 2.7 Å resolution Munshi et al. (2002), J. Biol. ...Chem.277, 38797–38802. In order to obtain a high‐resolution structure, a number of variants of IGFRK‐0P were prepared and screened for crystallization. A double mutant with E1067A and E1069A substitutions within the kinase‐insert region resulted in crystals that diffracted to 1.5 Å resolution. Overall, the structure of the mutant IGFRK‐0P is similar to that of the wild‐type IGFRK‐0P structure, with the exception of the previously disordered kinase‐insert region in the wild type having become fixed. In addition, amino‐acid residues 947–952 at the N‐terminus are well defined in the mutant structure. The monomeric protein structure is folded into two lobes connected by a hinge region, with the catalytic center situated at the interface of the two lobes. Two molecules of IGFRK‐0P in the asymmetric unit are associated as a dimer and two different types of dimers with their ATP‐binding clefts either facing towards or away from each other are observed. The current refined model consists of a dimer and 635 water molecules.
RORγt is critical for the differentiation and proliferation of Th17 cells associated with several chronic autoimmune diseases. We report the discovery of a novel allosteric binding site on the ...nuclear receptor RORγt. Co-crystallization of the ligand binding domain (LBD) of RORγt with a series of small-molecule antagonists demonstrates occupancy of a previously unreported allosteric binding pocket. Binding at this non-canonical site induces an unprecedented conformational reorientation of helix 12 in the RORγt LBD, which blocks cofactor binding. The functional consequence of this allosteric ligand-mediated conformation is inhibition of function as evidenced by both biochemical and cellular studies. RORγt function is thus antagonized in a manner molecularly distinct from that of previously described orthosteric RORγt ligands. This brings forward an approach to target RORγt for the treatment of Th17-mediated autoimmune diseases. The elucidation of an unprecedented modality of pharmacological antagonism establishes a mechanism for modulation of nuclear receptors.
As many processes in the preclinical drug discovery process become highly parallel, the need to also produce a large number of different proteins in parallel has become acute, such as for protein ...crystallization and activity screening. In turn, the requisite DNA constructions to produce these proteins must now be done at a rate that requires automated cloning procedures, each with an intrinsic low failure probability per sample. The high-throughput cloning solutions presented here achieve production of 192 different expression plasmids at a success rate of greater than 95% of the targeted open reading frames. Time for completion of the set by one person is reduced to approximately 11 working days, starting with polymerase chain reactions for a number of source clones and ending with purified expression plasmids. Achievement of this throughput utilizes the following: (1) the Beckman Coulter (Fullerton, CA) Biomek FX liquid handler for most manipulations, (2) Gateway cloning technology (Invitrogen Corp., Carlsbad, CA), and (3) computer programs designed for parallel processing of all sample information, including primer design and the resulting DNA and protein sequence assembly. Exemplary data are presented for discovery of a form of the Rho-kinase that crystallizes (ROCK2).
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