The CDK9 C-helix Exhibits Conformational Plasticity That May Explain the Selectivity of CAN508

CDK9 is the kinase of positive transcription elongation factor b and facilitates the transition of paused RNA polymerase II to processive transcription elongation. CDK9 is a validated target for the treatment of cancer, cardiac hypertrophy, and human immunodeficiency virus. Here we analyze different CDK9/cyclin T variants to identify a form of the complex amenable to use in inhibitor design. To demonstrate the utility of this system, we have determined the crystal structures of CDK9/cyclin T and CDK2/cyclin A bound to the CDK9-specific inhibitor CAN508. Comparison of the structures reveals CDK9-specific conformational changes and identifies a CDK9-specific hydrophobic pocket, adjacent to the αC-helix. By comparison with a previously published structure of CDK9/cyclin T/human immunodeficiency virus TAT we find that the CDK9 αC-helix has a degree of conformational variability that has the potential to be exploited for inhibitor design

X-ray diffraction images for an MDM2/Nutlin-3a complex

<p>This submission includes a zip archive of diffraction images recorded with the MARMOSAIC 225 mm CCD detector at the ESRF beam line ID23-2. Relevant meta data can be found in the headers of those diffraction images or in the Protein Data Bank entry 4HG7.</p

An Inhibitor’s-Eye View of the ATP-Binding Site of CDKs in Different Regulatory States

We have used a chemically diverse panel of kinase inhibitors to assess the chemical similarity of the ATP-binding sites of cyclin-dependent kinase (CDK) subfamily members in a range of activation states. Using this approach, we find that different activation states of a particular CDK may differ from each other as much as different CDKs in the same activation state. We also find that inhibitors discriminate more effectively among CDK family members in their monomeric state than in their cyclin-bound state, providing direct evidence for the belief that selective binding to inactive kinase states might be more readily achieved than selective binding to active states

Benzene Probes in Molecular Dynamics Simulations Reveal Novel Binding Sites for Ligand Design

Protein flexibility poses a major challenge in binding site identification. Several computational pocket detection methods that utilize small-molecule probes in molecular dynamics (MD) simulations have been developed to address this issue. Although they have proven hugely successful at reproducing experimental structural data, their ability to predict new binding sites that are yet to be identified and characterized has not been demonstrated. Here, we report the use of benzenes as probe molecules in ligand-mapping MD (LMMD) simulations to predict the existence of two novel binding sites on the surface of the oncoprotein MDM2. One of them was serendipitously confirmed by biophysical assays and X-ray crystallography to be important for the binding of a new family of hydrocarbon stapled peptides that were specifically designed to target the other putative site. These results highlight the predictive power of LMMD and suggest that predictions derived from LMMD simulations can serve as a reliable basis for the identification of novel ligand binding sites in structure-based drug design

8‑Substituted <i>O</i>⁶‑Cyclohexylmethylguanine CDK2 Inhibitors: Using Structure-Based Inhibitor Design to Optimize an Alternative Binding Mode

Evaluation of the effects of purine <i>C</i>-8 substitution within a series of CDK1/2-selective <i>O</i>⁶-cyclohexylmethylguanine derivatives revealed that potency decreases initially with increasing size of the alkyl substituent. Structural analysis showed that <i>C</i>-8 substitution is poorly tolerated, and to avoid unacceptable steric interactions, these compounds adopt novel binding modes. Thus, 2-amino-6-cyclohexylmethoxy-8-isopropyl-9<i>H</i>-purine adopts a “reverse” binding mode where the purine backbone has flipped 180°. This provided a novel lead chemotype from which we have designed more potent CDK2 inhibitors using, in the first instance, quantum mechanical energy calculations. Introduction of an <i>ortho</i>-tolyl or <i>ortho</i>-chlorophenyl group at the purine C-8 position restored the potency of these “reverse” binding mode inhibitors to that of the parent 2-amino-6-cyclohexylmethoxy-9<i>H</i>-purine. By contrast, the corresponding 8-(2-methyl-3-sulfamoylphenyl)-purine derivative exhibited submicromolar CDK2-inhibitory activity by virtue of engineered additional interactions with Asp86 and Lys89 in the reversed binding mode, as confirmed by X-ray crystallography

Kuntouttavan työtoiminnan prosessien kehittäminen

Tämän opinnäytetyön tarkoituksena oli kehittää kuntouttavan työtoiminnan prosesseja Pohjois-Satakunnan peruspalveluliikelaitoskuntayhtymässä. Kehittämistehtävässä kehitettiin toimivat mallit kuntouttavan työtoiminnan palveluun kunta -ja palvelurakenneuudistuksen mukanaan tuomassa uudessa organisaatiossa. Kuntouttavan työtoiminnan palvelua kehitettiin toimintatutkimuksen menetelmillä, jossa toimintatutkimuksen syklit, suunnittelu, toiminta, havainnointi ja reflektointi seurasivat toisiaan. Kehittämistehtävässä tarkasteltiin kuntouttavan työtoiminnan palvelua organisaation ja työntekijöiden näkökulmasta, mutta myös asiakasnäkökulma otettiin huomioon. Tutkimuksen aineistona käytettiin reflektoivaa päiväkirjaa sekä työntekijöiden ja asiakkaiden haastatteluja. Haastattelut tehtiin teemahaastattelun menetelmällä. Kehittämistehtävässä muodostettiin prosessikaaviot aktivointisuunnitelman laatimisesta, kuntouttavan työtoiminnan prosessista sekä työnjaosta eri toimipisteiden välillä. Lisäksi kehitettiin työntekijöiden työn yhtenäistämiseksi erilaisia malleja kuntouttavan työtoiminnan ohjaukseen. Kehittämistehtävän perusteella voidaan kuntayhtymän eri toimipisteissä tarjota tasavertaista ja yhdenmukaista palvelua ja asiakkaiden saatavilla ovat laadukkaat ja kattavat kuntouttavan työtoiminnan palvelut. Kehittämistehtävän mukaan tulevaisuuden haasteena on kuntouttavan työtoiminnan asiakasmäärien kasvu, osaamista vastaavien työtoimintapaikkojen löytyminen sekä riittävä ohjausresurssi, myös aktiivisen sosiaalipolitiikan mukanaan tuomat lakimuutokset tuovat haastetta kuntouttavan työtoiminnan järjestämiseen. Kehittämisen keskeisinä käsitteinä olivat kunta- ja palvelurakenneuudistus, aktiivinen sosiaalipolitiikka, kuntouttava työtoiminta, osallisuus ja kuntouttavan työtoiminnan vaikuttavuus Avainsanat: sosiaalipolitiikka, työllisyyspolitiikka, työttömät, kuntouttava työtoiminta, palvelurakenne.The aim of this development project was to improve rehabilitative employment activities and working processes. This development project was executed in the new organization of Social and Healthcare federation of municipalities in Pohjois-Satakunta. The rehabilitative employment activity and its processes were improved with methods of activity analysis. There planning, action, observation and reflection had been followed by each other. The aim of the rehabilitative employment activity is working life or maintaining the unemployed person’s ability to work. The materials of this developments project were collected with interviews and reflective diary. During this development project had been created new patterns how to guide new clients to the rehabilitative employment activities. This development project is also description of that process. Keywords: rehabilitative employment activity, activation policy, unemployment, social politics