In Silico Investigation of Potential Applications of Gamma Carbonic Anhydrases as Catalysts of CO2 Biomineralization Processes: A Visit to the Thermophilic Bacteria Persephonella hydrogeniphila,Persephonella marina,Thermosulfidibacter takaii,and Thermus thermophilus

Carbonic anhydrases (CAs) have been identified as ideal catalysts for CO₂ sequestration. Here, we report the sequence and structural analyses as well as the molecular dynamics (MD) simulations of four γ-CAs from thermophilic bacteria. Three of these, Persephonella marina, Persephonella hydrogeniphila, and Thermosulfidibacter takaii originate from hydrothermal vents and one, Thermus thermophilus HB8, from hot springs. Protein sequences were retrieved and aligned with previously characterized γ-CAs, revealing differences in the catalytic pocket residues. Further analysis of the structures following homology modeling revealed a hydrophobic patch in the catalytic pocket, presumed important for CO₂ binding. Monitoring of proton shuttling residue His69 (P. marina γ-CA numbering) during MD simulations of P. hydrogeniphila and P. marina’s γ-CAs (γ-PhCA and γ-PmCA), showed a different behavior to that observed in the γ-CA of Escherichia coli, which periodically coordinates Zn²⁺. This work also involved the search for hotspot residues that contribute to interface stability. Some of these residues were further identified as key in protein communication via betweenness centrality metric of dynamic residue network analysis. T. takaii’s γ-CA showed marginally lower thermostability compared to the other three γ-CA proteins with an increase in conformations visited at high temperatures being observed. Hydrogen bond analysis revealed important interactions, some unique and others common in all γ-CAs, which contribute to interface formation and thermostability. The seemingly thermostable γ-CA from T. thermophilus strangely showed increased unsynchronized residue motions at 423 K. γ-PhCA and γ-PmCA were, however, preliminarily considered suitable as prospective thermostable CO₂ sequestration agents.     

Heterogeneity in Youth Symptom Trajectories Following Psychiatric Crisis: Predictors and Placement Outcome.

The authors examined heterogeneity in symptom trajectories among youths following psychiatric crises as well as the psychosocial correlates and placement outcomes associated with identified trajectories. Using semiparametric mixture modeling with 156 youths approved for psychiatric hospitalization, the authors identified 5 trajectories based on symptoms over the 16 months following crisis: high improved, high unimproved, borderline improved, borderline unimproved, and subclinical. Membership in unimproved symptom groups was associated with less suicidality, younger age, more youth hopelessness, and more caregiver empowerment. Improved symptom group membership predicted long-term decreases in days in out-of-home placements. More important, and in contrast with general impressions from the existing literature, findings suggest that a substantive proportion of youths with serious emotional disturbance sustain high levels of symptomatology following intensive mental health services. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

The likelihood of violent behaviour: Predictions, postdictions, and hindsight bias.

Studied the effects of variations in base rate (BR) information and temporal context on predictions of violence and hindsight bias (HB). 270 undergraduates estimated the likelihood of violent offenders committing another violent offense based upon fictitious case history information. Ss were given 1 of 2 population BRs of the likelihood of reoffending (30% or 70%) and a recidivism estimation question that was phrased either predictively, predictively with a given outcome of either recidivism or no recidivism, or postdictively. Although different offenders were perceived as differentially likely to reoffend, estimated likelihood of recidivism was affected by neither BR nor temporal phrasing of the estimation task. A 2nd study of 182 undergraduates, using shorter case histories, also found no BR effect. HB effects were weak and inconsistent in both studies. (French abstract) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Behavioral model synthesis with Cones

The Cones synthesis system for automatic generation of VLSI implementations is discussed. Named for the cones in sequential logic, Cones takes behavioral models written in C and produces gate-level implementations in technologies such as standard cells and programmable logic arrays or programmable logic devices. The overall design is produced faster, more efficiently, and with fewer errors. Designers are free to concentrate on functions, instead of on the details of the implementation technology     

The additivity of substrate fragments in enzyme-ligand binding

BACKGROUND: Enzymes have evolved to recognise their target substrates with exquisite selectivity and specificity. Whether fragments of the substrate--perhaps never available to the evolving enzyme--are bound in the same manner as the parent substrate addresses the fundamental basis of specificity. An understanding of the relative contributions of individual portions of ligand molecules to the enzyme-binding interaction may offer considerable insight into the principles of substrate recognition. RESULTS: We report 12 crystal structures of Escherichia coli thymidylate synthase in complexes with available fragments of the substrate (dUMP), both with and without the presence of a cofactor analogue. The structures display considerable fidelity of binding mode and interactions. These complexes reveal several interesting features: the cofactor analogue enhances the localisation of substrate and substrate fragments near the reactive thiol; the ribose moiety reduces local disorder through additional specific enzyme-ligand interactions; the pyrimidine has multiple roles, ranging from stereospecificity to mechanistic competence; and the glycosidic linkage has an important role in the formation of a covalent attachment between substrate and enzyme. CONCLUSIONS: The requirements of ligand-protein binding can be understood in terms of the binding of separate fragments of the ligand. Fragments which are subsystems of the natural substrate for the enzyme confer specific contributions to the binding affinity, orientation or electrostatics of the enzymatic mechanism. This ligand-binding analysis provides a complementary method to the more prevalent approaches utilising site-directed mutagenesis. In addition, these observations suggest a modular approach for rational drug design utilising chemical fragments.     

Over-production of 5-enolpyruvylshikimate-3-phosphate synthase in Escherichia coli: use of the T7 promoter

5-Enolpyruvylshikimate-3-phosphate (EPSP) synthase, the productof the Escherichia coli aroA gene, has been overproduced inE.coli BL21(DE3) under the control of the T7 gene 10 promoterand ribosome binding site, to a level of {small tilde}50% oftotal cell protein. EPSP synthase is the primary target of thepost-emergence herbicide, glyphosate, commonly known as Roundup^TM.A simple two step purification is described, which results in99% pure homogeneous protein (as determined by PAGE). The integrityof the protein has been compared with previously characterizedprotein from .E.coli AB2829(pKD501) by determination of itskinetic parameters, N-terminal protein and DNA sequences, aminoacid analysis and ¹³C-NMR spectroscopy. This new overproducingstrain readily provides the gram quantities of highly pure proteinrequired for NMR studies of the active site and the developmentof novel time-resolved solid-state NMR techniques currentlyunderway in this laboratory.     

Torrefied agro-industrial residue as filler in natural rubber compounds

This study explored the feasibility of using torrefied biomass as a reinforcing filler in natural rubber compounds. Carbon black was then replaced with the torrefied biomass in elastomer formulations for concentrations varying from 0% to 100% (60 parts per hundred rubber or phr total). Their influence on the curing process, dynamic properties, and mechanical properties was investigated. Results were compared with the properties of vulcanizates containing solely carbon black fillers. Time to cure (t₉₀) for compounds with torrefied biomass fillers increased, while filler-filler interactions (ΔG') decreased, compared to carbon black controls. At low strains, the tan δ values of the torrefied fillers vulcanizates were similar to the controls. Incorporation of torrefied biomass into natural rubber decreased compound tensile strength and modulus but increased elongation. Replacement with torrefied fillers resulted in a weaker filler network in the matrix. Still, results showed that moderate substitution concentrations (~20 phr) could be feasible for some natural rubber applications.     

Case-Based Learning: Predictive Features in Indexing

Interest in psychological experimentation from the Artificial Intelligence community often takes the form of rigorous post-hoc evaluation of completed computer models. Through an example of our own collaborative research, we advocate a different view of how psychology and AI may be mutually relevant, and propose an integrated approach to the study of learning in humans and machines. We begin with the problem of learning appropriate indices for storing and retrieving information from memory. From a planning task perspective, the most useful indices may be those that predict potential problems and access relevant plans in memory, improving the planner's ability to predict and avoid planning failures. This predictive features hypothesis is then supported as a psychological claim, with results showing that such features offer an advantage in terms of the selectivity of reminding because they more distinctively characterize planning situations where differing plans are appropriate.We present a specific case-based model of plan execution, RUNNER, along with its indices for recognizing when to select particular plans—appropriateness conditions—and how these predictive indices serve to enhance learning. We then discuss how this predictive features claim as implemented in the RUNNER model is then tested in a second set of psychological studies. The results show that learning appropriateness conditions results in greater success in recognizing when a past plan is in fact relevant in current processing, and produces more reliable recall of the related information. This form of collaboration has resulted in a unique integration of computational and empirical efforts to create a model of case-based learning.     

Domain flexibility in retroviral proteases: structural implications for drug resistant mutations

Rigid body rotation of five domains and movements within their interfacial joints provide a rational context for understanding why HIV protease mutations that arise in drug resistant strains are often spatially removed from the drug or substrate binding sites. Domain motions associated with substrate binding in the retroviral HIV-1 and SIV proteases are identified and characterized. These motions are in addition to closure of the flaps and result from rotations of approximately 6-7 degrees at primarily hydrophobic interfaces. A crystal structure of unliganded SIV protease (incorporating the point mutation Ser 4 His to stabilize the protease against autolysis) was determined to 2.0 A resolution in a new space group, P3221. The structure is in the most "open" conformation of any retroviral protease so far examined, with six residues of the flaps disordered. Comparison of this and unliganded HIV structures, with their respective liganded structures by difference distance matrixes identifies five domains of the protease dimer that move as rigid bodies against one another: one terminal domain encompassing the N- and C-terminal beta sheet of the dimer, two core domains containing the catalytic aspartic acids, and two flap domains. The two core domains rotate toward each other on substrate binding, reshaping the binding pocket. We therefore show that, for enzymes, mutations at interdomain interfaces that favor the unliganded form of the target active site will increase the off-rate of the inhibitor, allowing the substrate greater access for catalysis. This offers a mechanism of resistance to competitive inhibitors, especially when the forward enzymatic reaction rate exceeds the rate of substrate dissociation.