Coot is a molecular-graphics application for model building and validation of biological macromolecules. The program displays electron-density maps and atomic models and allows model manipulations such as idealization, real-space refinement, manual rotation/translation, rigid-body fitting, ligand search, solvation, mutations, rotamers and Ramachandran idealization. Furthermore, tools are provided for model validation as well as interfaces to external programs for refinement, validation and graphics. The software is designed to be easy to learn for novice users, which is achieved by ensuring that tools for common tasks are `discoverable' through familiar user-interface elements (menus and toolbars) or by intuitive behaviour (mouse controls). Recent developments have focused on providing tools for expert users, with customisable key bindings, extensions and an extensive scripting interface. The software is under rapid development, but has already achieved very widespread use within the crystallographic community. The current state of the software is presented, with a description of the facilities available and of some of the underlying methods employed.

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Features and development of Coot.

J. Appl. Cryst.の発刊に際して

A newly described coronavirus named severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which is the causative agent of coronavirus disease 2019 (COVID-19), has infected over 2.3 million people, led to the death of more than 160,000 individuals and caused worldwide social and economic disruption1,2. There are no antiviral drugs with proven clinical efficacy for the treatment of COVID-19, nor are there any vaccines that prevent infection with SARS-CoV-2, and efforts to develop drugs and vaccines are hampered by the limited knowledge of the molecular details of how SARS-CoV-2 infects cells. Here we cloned, tagged and expressed 26 of the 29 SARS-CoV-2 proteins in human cells and identified the human proteins that physically associated with each of the SARS-CoV-2 proteins using affinity-purification mass spectrometry, identifying 332 high-confidence protein–protein interactions between SARS-CoV-2 and human proteins. Among these, we identify 66 druggable human proteins or host factors targeted by 69 compounds (of which, 29 drugs are approved by the US Food and Drug Administration, 12 are in clinical trials and 28 are preclinical compounds). We screened a subset of these in multiple viral assays and found two sets of pharmacological agents that displayed antiviral activity: inhibitors of mRNA translation and predicted regulators of the sigma-1 and sigma-2 receptors. Further studies of these host-factor-targeting agents, including their combination with drugs that directly target viral enzymes, could lead to a therapeutic regimen to treat COVID-19. A human–SARS-CoV-2 protein interaction map highlights cellular processes that are hijacked by the virus and that can be targeted by existing drugs, including inhibitors of mRNA translation and predicted regulators of the sigma receptors.

/pdf/a-sars-cov-2-protein-interaction-map-reveals-targets-for-5d1m2n6hc1.pdf

A SARS-CoV-2 protein interaction map reveals targets for drug repurposing.

Microbe-associated molecular patterns (MAMPs) are molecular signatures typical of whole classes of microbes, and their recognition plays a key role in innate immunity. Endogenous elicitors are similarly recognized as damage-associated molecular patterns (DAMPs). This review focuses on the diversity of MAMPs/DAMPs and on progress to identify the corresponding pattern recognition receptors (PRRs) in plants. The two best-characterized MAMP/PRR pairs, flagellin/FLS2 and EF-Tu/EFR, are discussed in detail and put into a phylogenetic perspective. Both FLS2 and EFR are leucine-rich repeat receptor kinases (LRR-RKs). Upon treatment with flagellin, FLS2 forms a heteromeric complex with BAK1, an LRR-RK that also acts as coreceptor for the brassinolide receptor BRI1. The importance of MAMP/PRR signaling for plant immunity is highlighted by the finding that plant pathogens use effectors to inhibit PRR complexes or downstream signaling events. Current evidence indicates that MAMPs, DAMPs, and effectors are all perceived as danger signals and induce a stereotypic defense response.

A Renaissance of Elicitors: Perception of Microbe-Associated Molecular Patterns and Danger Signals by Pattern-Recognition Receptors

Plants are engaged in a continuous co-evolutionary struggle for dominance with their pathogens. The outcomes of these interactions are of particular importance to human activities, as they can have dramatic effects on agricultural systems. The recent convergence of molecular studies of plant immunity and pathogen infection strategies is revealing an integrated picture of the plant-pathogen interaction from the perspective of both organisms. Plants have an amazing capacity to recognize pathogens through strategies involving both conserved and variable pathogen elicitors, and pathogens manipulate the defence response through secretion of virulence effector molecules. These insights suggest novel biotechnological approaches to crop protection.

/pdf/plant-immunity-towards-an-integrated-view-of-plant-pathogen-3835470zf1.pdf

Plant immunity: towards an integrated view of plant―pathogen interactions

Silent information regulator 2 (Sir2) proteins (sirtuins) are nicotinamide adenine dinucleotide-dependent deacetylases that regulate important biological processes. Mammals have seven sirtuins, Sirt1 to Sirt7. Four of them (Sirt4 to Sirt7) have no detectable or very weak deacetylase activity. We found that Sirt5 is an efficient protein lysine desuccinylase and demalonylase in vitro. The preference for succinyl and malonyl groups was explained by the presence of an arginine residue (Arg(105)) and tyrosine residue (Tyr(102)) in the acyl pocket of Sirt5. Several mammalian proteins were identified with mass spectrometry to have succinyl or malonyl lysine modifications. Deletion of Sirt5 in mice appeared to increase the level of succinylation on carbamoyl phosphate synthase 1, which is a known target of Sirt5. Thus, protein lysine succinylation may represent a posttranslational modification that can be reversed by Sirt5 in vivo.

/pdf/sirt5-is-a-nad-dependent-protein-lysine-demalonylase-and-3765xep097.pdf

Sirt5 is a NAD-dependent protein lysine demalonylase and desuccinylase.

The Sir2 family of enzymes or sirtuins are known as nicotinamide adenine dinucleotide (NAD)-dependent deacetylases and have been implicated in the regulation of transcription, genome stability, metabolism and lifespan. However, four of the seven mammalian sirtuins have very weak deacetylase activity in vitro. Here we show that human SIRT6 efficiently removes long-chain fatty acyl groups, such as myristoyl, from lysine residues. The crystal structure of SIRT6 reveals a large hydrophobic pocket that can accommodate long-chain fatty acyl groups. We demonstrate further that SIRT6 promotes the secretion of tumour necrosis factor-α (TNF-α) by removing the fatty acyl modification on K19 and K20 of TNF-α. Protein lysine fatty acylation has been known to occur in mammalian cells, but the function and regulatory mechanisms of this modification were unknown. Our data indicate that protein lysine fatty acylation is a novel mechanism that regulates protein secretion. The discovery of SIRT6 as an enzyme that controls protein lysine fatty acylation provides new opportunities to investigate the physiological function of a protein post-translational modification that has been little studied until now.

/pdf/sirt6-regulates-tnf-a-secretion-through-hydrolysis-of-long-27xhhtr1hs.pdf

SIRT6 regulates TNF-α secretion through hydrolysis of long-chain fatty acyl lysine

Sirt6 Regulates Tnf-Alpha Secretion Through Hydrolysis of Long-Chain Fatty Acyl Lysine

Metallo-β-lactamases (MBLs) hydrolyze most β-lactam antibiotics, and bacteria containing this kind of enzyme pose a serious threat to the public health. The newly identified New Delhi MBL (NDM-1) is a new member of this family that shows tight binding to penicillin and cephalosporins. The rapid dissemination of NDM-1 in clinically relevant bacteria has become a global concern. However, no clinically useful inhibitors against MBLs exist, partly due to the lack of knowledge about the catalysis mechanism of this kind of enzyme. Here we report the crystal structure of this novel enzyme in complex with a hydrolyzed ampicillin at its active site at 1.3-A resolution. Structural comparison with other MBLs revealed a new hydrolysis mechanism applicable to all three subclasses of MBLs, which might help the design of mechanism based inhibitors.

Crystal structure of NDM-1 reveals a common β-lactam hydrolysis mechanism

Interplay among four genes—egl-1, ced-9, ced-4 and ced-3—controls the onset of programmed cell death in the nematode Caenorhabditis elegans. Activation of the cell-killing protease CED-3 requires CED-4. However, CED-4 is constitutively inhibited by CED-9 until its release by EGL-1. Here we report the crystal structure of the CED-4–CED-9 complex at 2.6 A resolution, and a complete reconstitution of the CED-3 activation pathway using homogeneous proteins of CED-4, CED-9 and EGL-1. One molecule of CED-9 binds to an asymmetric dimer of CED-4, but specifically recognizes only one of the two CED-4 molecules. This specific interaction prevents CED-4 from activating CED-3. EGL-1 binding induces pronounced conformational changes in CED-9 that result in the dissociation of the CED-4 dimer from CED-9. The released CED-4 dimer further dimerizes to form a tetramer, which facilitates the autoactivation of CED-3. Together, our studies provide important insights into the regulation of cell death activation in C. elegans.

Quan Hao

Papers

Sirt5 is a NAD-dependent protein lysine demalonylase and desuccinylase.

SIRT6 regulates TNF-α secretion through hydrolysis of long-chain fatty acyl lysine

Sirt6 Regulates Tnf-Alpha Secretion Through Hydrolysis of Long-Chain Fatty Acyl Lysine

Crystal structure of NDM-1 reveals a common β-lactam hydrolysis mechanism

Structure of the CED-4–CED-9 complex provides insights into programmed cell death in Caenorhabditis elegans