Publications by the Blundell group

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Arpeggio: A Web Server for Calculating and Visualising Interatomic Interactions in Protein Structures.

Mon, 14/08/2017 - 13:39
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Arpeggio: A Web Server for Calculating and Visualising Interatomic Interactions in Protein Structures.

J Mol Biol. 2017 Feb 03;429(3):365-371

Authors: Jubb HC, Higueruelo AP, Ochoa-Montaño B, Pitt WR, Ascher DB, Blundell TL

Abstract
Interactions between proteins and their ligands, such as small molecules, other proteins, and DNA, depend on specific interatomic interactions that can be classified on the basis of atom type and distance and angle constraints. Visualisation of these interactions provides insights into the nature of molecular recognition events and has practical uses in guiding drug design and understanding the structural and functional impacts of mutations. We present Arpeggio, a web server for calculating interactions within and between proteins and protein, DNA, or small-molecule ligands, including van der Waals', ionic, carbonyl, metal, hydrophobic, and halogen bond contacts, and hydrogen bonds and specific atom-aromatic ring (cation-π, donor-π, halogen-π, and carbon-π) and aromatic ring-aromatic ring (π-π) interactions, within user-submitted macromolecule structures. PyMOL session files can be downloaded, allowing high-quality publication images of the interactions to be generated. Arpeggio is implemented in Python and available as a user-friendly web interface at http://structure.bioc.cam.ac.uk/arpeggio/ and as a downloadable package at https://bitbucket.org/harryjubb/arpeggio.

PMID: 27964945 [PubMed - indexed for MEDLINE]

Categories: Publications

DNA-PKcs, Allostery, and DNA Double-Strand Break Repair: Defining the Structure and Setting the Stage.

Mon, 31/07/2017 - 11:15

DNA-PKcs, Allostery, and DNA Double-Strand Break Repair: Defining the Structure and Setting the Stage.

Methods Enzymol. 2017;592:145-157

Authors: Chirgadze DY, Ascher DB, Blundell TL, Sibanda BL

Abstract
DNA-dependent protein kinase catalytic subunit (DNA-PKcs) is central to the regulation of the DNA damage response and repair through nonhomologous end joining. The structure has proved challenging due to its large size and multiple HEAT repeats. We have recently reported crystals of selenomethionine-labeled DNA-PKcs complexed with native KU80ct194 (KU80 residues 539-732) diffracting to 4.3Å resolution. The novel use of crystals of selenomethionine-labeled protein expressed in HeLa cells has facilitated the use of single anomalous X-ray scattering of this 4128 amino acid, multiple HEAT-repeat structure. The monitoring of the selenomethionines in the anomalous-difference density map has allowed the checking of the amino acid residue registration in the electron density, and the labeling of the Ku-C-terminal moiety with selenomethionine has further allowed its identification in the structure of the complex with DNA-PKcs. The crystal structure defines a stage on which many of the components assemble and regulate the kinase activity through modulating the conformation and allosteric regulation of kinase activity.

PMID: 28668119 [PubMed - in process]

Categories: Publications

Structural biology and the design of new therapeutics: from HIV and cancer to mycobacterial infections.

Mon, 31/07/2017 - 11:15
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Structural biology and the design of new therapeutics: from HIV and cancer to mycobacterial infections.

J Mol Biol. 2017 Jun 22;:

Authors: Thomas SE, Mendes V, Kim SY, Malhotra S, Ochoa-Montaño B, Blaszczyk M, Blundell TL

Abstract
Interest in applications of protein crystallography to medicine was evident as the first high-resolution structures emerged in the 50s and 60s. In Cambridge Max Perutz and John Kendrew sought to understand mutations in sickle cell and other genetic diseases related to haemoglobin, while in Oxford the group of Dorothy Hodgkin became interested in long-lasting zinc-insulin crystals for treatment of diabetes and later considered insulin redesign as synthetic insulins became possible. The use of protein crystallography in structure-guided drug discovery emerged as enzyme structures allowed the identification of potential inhibitor-binding sites and optimisation of interactions of hits using the structure of the target protein. Early examples of this approach were the use of the structure of renin to design anti-hypertensives and the structure of HIV protease in design of AIDS antivirals. More recently, use of structure-guided design with fragment-based drug discovery, which reduces the size of screening libraries by decreasing complexity, has improved ligand efficiency in drug design and has been used to progress three oncology drugs through clinical trials to FDA approval. We exemplify current developments in structure-guided target identification and fragment-based lead discovery with efforts to develop new antimicrobials for mycobacterial infections.

PMID: 28648615 [PubMed - as supplied by publisher]

Categories: Publications

Engineering Archeal Surrogate Systems for the Development of Protein-Protein Interaction Inhibitors against Human RAD51.

Mon, 31/07/2017 - 11:15
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Engineering Archeal Surrogate Systems for the Development of Protein-Protein Interaction Inhibitors against Human RAD51.

J Mol Biol. 2016 Nov 20;428(23):4589-4607

Authors: Moschetti T, Sharpe T, Fischer G, Marsh ME, Ng HK, Morgan M, Scott DE, Blundell TL, R Venkitaraman A, Skidmore J, Abell C, Hyvönen M

Abstract
Protein-protein interactions (PPIs) are increasingly important targets for drug discovery. Efficient fragment-based drug discovery approaches to tackle PPIs are often stymied by difficulties in the production of stable, unliganded target proteins. Here, we report an approach that exploits protein engineering to "humanise" thermophilic archeal surrogate proteins as targets for small-molecule inhibitor discovery and to exemplify this approach in the development of inhibitors against the PPI between the recombinase RAD51 and tumour suppressor BRCA2. As human RAD51 has proved impossible to produce in a form that is compatible with the requirements of fragment-based drug discovery, we have developed a surrogate protein system using RadA from Pyrococcus furiosus. Using a monomerised RadA as our starting point, we have adopted two parallel and mutually instructive approaches to mimic the human enzyme: firstly by mutating RadA to increase sequence identity with RAD51 in the BRC repeat binding sites, and secondly by generating a chimeric archaeal human protein. Both approaches generate proteins that interact with a fourth BRC repeat with affinity and stoichiometry comparable to human RAD51. Stepwise humanisation has also allowed us to elucidate the determinants of RAD51 binding to BRC repeats and the contributions of key interacting residues to this interaction. These surrogate proteins have enabled the development of biochemical and biophysical assays in our ongoing fragment-based small-molecule inhibitor programme and they have allowed us to determine hundreds of liganded structures in support of our structure-guided design process, demonstrating the feasibility and advantages of using archeal surrogates to overcome difficulties in handling human proteins.

PMID: 27725183 [PubMed - indexed for MEDLINE]

Categories: Publications

Identifying Interactions that Determine Fragment Binding at Protein Hotspots.

Mon, 31/07/2017 - 11:15
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Identifying Interactions that Determine Fragment Binding at Protein Hotspots.

J Med Chem. 2016 May 12;59(9):4314-25

Authors: Radoux CJ, Olsson TS, Pitt WR, Groom CR, Blundell TL

Abstract
Locating a ligand-binding site is an important first step in structure-guided drug discovery, but current methods do little to suggest which interactions within a pocket are the most important for binding. Here we illustrate a method that samples atomic hotspots with simple molecular probes to produce fragment hotspot maps. These maps specifically highlight fragment-binding sites and their corresponding pharmacophores. For ligand-bound structures, they provide an intuitive visual guide within the binding site, directing medicinal chemists where to grow the molecule and alerting them to suboptimal interactions within the original hit. The fragment hotspot map calculation is validated using experimental binding positions of 21 fragments and subsequent lead molecules. The ligands are found in high scoring areas of the fragment hotspot maps, with fragment atoms having a median percentage rank of 97%. Protein kinase B and pantothenate synthetase are examined in detail. In each case, the fragment hotspot maps are able to rationalize a Free-Wilson analysis of SAR data from a fragment-based drug design project.

PMID: 27043011 [PubMed - indexed for MEDLINE]

Categories: Publications

Virtual Screening and X-ray Crystallography Identify Non-Substrate Analog Inhibitors of Flavin-Dependent Thymidylate Synthase.

Mon, 26/06/2017 - 15:18
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Virtual Screening and X-ray Crystallography Identify Non-Substrate Analog Inhibitors of Flavin-Dependent Thymidylate Synthase.

J Med Chem. 2016 Oct 13;59(19):9269-9275

Authors: Luciani R, Saxena P, Surade S, Santucci M, Venturelli A, Borsari C, Marverti G, Ponterini G, Ferrari S, Blundell TL, Costi MP

Abstract
Thymidylate synthase X (ThyX) represents an attractive target for tuberculosis drug discovery. Herein, we selected 16 compounds through a virtual screening approach. We solved the first X-ray crystal structure of Thermatoga maritima (Tm) ThyX in complex with a nonsubstrate analog inhibitor. Given the active site similarities between Mycobacterium tuberculosis ThyX (Mtb-ThyX) and Tm-ThyX, our crystal structure paves the way for a structure-based design of novel antimycobacterial compounds. The 1H-imidazo[4,5-d]pyridazine was identified as scaffold for the development of Mtb-ThyX inhibitors.

PMID: 27589670 [PubMed - indexed for MEDLINE]

Categories: Publications