Experiments with a Convex Polyhedral Analysis Tool for Logic Programs

Convex polyhedral abstractions of logic programs have been found very useful in deriving numeric relationships between program arguments in order to prove program properties and in other areas such as termination and complexity analysis. We present a tool for constructing polyhedral analyses of (constraint) logic programs. The aim of the tool is to make available, with a convenient interface, state-of-the-art techniques for polyhedral analysis such as delayed widening, narrowing, "widening up-to", and enhanced automatic selection of widening points. The tool is accessible on the web, permits user programs to be uploaded and analysed, and is integrated with related program transformations such as size abstractions and query-answer transformation. We then report some experiments using the tool, showing how it can be conveniently used to analyse transition systems arising from models of embedded systems, and an emulator for a PIC microcontroller which is used for example in wearable computing systems. We discuss issues including scalability, tradeoffs of precision and computation time, and other program transformations that can enhance the results of analysis.Comment: Paper presented at the 17th Workshop on Logic-based Methods in Programming Environments (WLPE2007

Constraint-Based Abstraction of a Model Checker for Infinite State Systems

Abstract. Abstract interpretation-based model checking provides an approach to verifying properties of infinite-state systems. In practice, most previous work on abstract model checking is either restricted to verifying universal properties, or develops special techniques for temporal logics such as modal transition sys-tems or other dual transition systems. By contrast we apply completely standard techniques for constructing abstract interpretations to the abstraction of a CTL semantic function, without restricting the kind of properties that can be verified. Furthermore we show that this leads directly to implementation of abstract model checking algorithms for abstract domains based on constraints, making use of an SMT solver.

Identification and characterization of <i>Helicobacter pylori</i> O-acetylserine-dependent cystathionine β-synthase, a distinct member of the PLP-II family

O-acetylserine sulfhydrylase (OASS) and cystathionine β-synthase (CBS) are members of the PLP-II family, and involved in L-cysteine production. OASS produces L-cysteine via a de novo pathway while CBS participates in the reverse transsulfuration pathway. O-acetylserine-dependent CBS (OCBS) was previously identified as a new member of the PLP-II family, which are predominantly seen in bacteria. The bacterium Helicobacter pylori possess only one OASS (hp0107) gene and we showed that the protein coded by this gene actually functions as an OCBS and utilizes L-homocysteine and O-acetylserine (OAS) to produce cystathionine. HpOCBS did not show CBS activity with the substrate L-serine and required OAS exclusively. The HpOCBS structure in complex with methionine showed a closed cleft state, explaining the initial mode of substrate binding. Sequence and structural analyses showed differences between the active sites of OCBS and CBS, and explain their different substrate preferences. We identified three hydrophobic residues near the active site of OCBS, corresponding to one serine and two tyrosine residues in CBSs. Mutational studies were performed on HpOCBS and Saccharomyces cerevisiae CBS. A ScCBS double mutant (Y158F/Y226V) did not display activity with L-serine, indicating indispensability of these polar residues for selecting substrate L-serine, however, did show activity with OAS.</p

Carbonic anhydrase II based biosensing of carbon dioxide at high temperature: an analytical and MD simulation study

Concentration of carbon dioxide (CO2) in the atmosphere has increased significantly due to anthropogenic activities and attributed as a major factor to global warming. Its detection by biosensing methods will provide an alternative for the assessment of CO2 concentration. Biomineralization of CO2 is one of the available methods for the biological conversion of CO2 to carbonate using a highly active enzyme, carbonic anhydrase II (CAII). CAII was used for the carbonation reaction to convert CO2 to CaCO3. The precipitation of calcium carbonate (CaCO3) was promoted in the presence of the CAII at 325 K. CAII showed an enhanced formation of solid CaCO3 through the acceleration of CO2 hydration rate at 325 K. Furthermore, the electrocatalytic properties of glassy carbon electrode enable us to determine the reduction peak potential values of CO2 through cyclic voltammetry at –1.75 and 0.3 V at 325 K. Molecular dynamic (MD) simulations were performed each at 50 ns time scale provided a deeper insight into the molecular basis of the CAII interaction with CO2 at different temperatures, highlighted that the CAII can detect CO2 up to 325 K. We assume that CAII could be an effective and economical biosensor for biomineralization of CO2 at high temperature 325 K

Structures of smooth muscle myosin and heavy meromyosin in the folded, shutdown state

Remodelling of the contractile apparatus within smooth muscle cells is an essential process that allows effective contractile activity over a wide range of cell lengths. The thick filaments may be redistributed via depolymerisation into inactive myosin monomers that have been detected in vitro, in which the long tail has a folded conformation. The structure of this folded molecule has been controversial. Using negative stain electron microscopy of individual folded molecules from turkey gizzard we show they are more compact than previously described, with heads and the three segments of the folded tail closely packed. Smooth muscle heavy meromyosin (HMM), which lacks two-thirds of the tail, closely resembles the equivalent parts of whole myosin. Image processing reveals a characteristic head region morphology for both HMM and myosin whose features are identifiable by comparison with less compact molecules. The two heads associate asymmetrically: the tip of one motor domain touches the base of the other, resembling the blocked and free heads of this HMM when it forms 2-D crystals on lipid. The tail of HMM lies between the heads, contacting the blocked motor domain, unlike in the 2-D crystal. The tail of the intact myosin is bent sharply and consistently at two positions close to residues 1175 and 1535. The first bend position correlates with a skip in the coiled coil sequence, the second does not. The first segment runs between the heads from the head-tail junction. Unexpectedly, the other segments associate only with the blocked head rather than both heads, such that the second bend lies at a specific position near the C-lobe of the blocked head regulatory light chain. Quantitative analysis of tail flexibility shows that the single coiled coil of HMM has an apparent Young’s modulus of about 0.5 GPa. The folded tail of the intact molecule is less flexible indicating interactions between the segments. The folded tail does not modify the compact head arrangement but stabilises it, indicating a structural mechanism for the very low ATPase activity of the folded molecule

Flexibility within the Heads of Muscle Myosin-2 Molecules

We show that negative-stain electron microscopy and image processing of nucleotide-free (apo) striated muscle myosin-2 subfragment-1 (S1), possessing one light chain or both light chains, is capable of resolving significant amounts of structural detail. The overall appearance of the motor and the lever is similar in rabbit, scallop and chicken S1. Projection matching of class averages of the different S1 types to projection views of two different crystal structures of apo S1 shows that all types most commonly closely resemble the appearance of the scallop S1 structure rather than the methylated chicken S1 structure. Methylation of chicken S1 has no effect on the structure of the molecule at this resolution: it too resembles the scallop S1 crystal structure. The lever is found to vary in its angle of attachment to the motor domain, with a hinge point located in the so-called pliant region between the converter and the essential light chain. The chicken S1 crystal structure lies near one end of the range of flexion observed. The Gaussian spread of angles of flexion suggests that flexibility is driven thermally, from which a torsional spring constant of ~ 23 pN·nm/rad2 is estimated on average for all S1 types, similar to myosin-5. This translates to apparent cantilever-type stiffness at the tip of the lever of 0.37 pN/nm. Because this stiffness is lower than recent estimates from myosin-2 heads attached to actin, we suggest that binding to actin leads to an allosteric stiffening of the motor–lever junction

Investigations on Binding Pattern of Kinase Inhibitors with PPAR γ

Peroxisome proliferator-activated receptor gamma (PPARγ) is a potential target for the treatment of several disorders. In view of several FDA approved kinase inhibitors, in the current study, we have investigated the interaction of selected kinase inhibitors with PPARγ using computational modeling, docking, and molecular dynamics simulations (MDS). The docked conformations and MDS studies suggest that the selected KIs interact with PPARγ in the ligand binding domain (LBD) with high positive predictive values. Hence, we have for the first time shown the plausible binding of KIs in the PPARγ ligand binding site. The results obtained from these in silico investigations warrant further evaluation of kinase inhibitors as PPARγ ligands in vitro and in vivo

Actomyosin Interaction: Mechanical and Energetic Properties in Different Nucleotide Binding States

The mechanics of the actomyosin interaction is central in muscle contraction and intracellular trafficking. A better understanding of the events occurring in the actomyosin complex requires the examination of all nucleotide-dependent states and of the energetic features associated with the dynamics of the cross-bridge cycle. The aim of the present study is to estimate the interaction strength between myosin in nucleotide-free, ATP, ADP·Pi and ADP states and actin monomer. The molecular models of the complexes were constructed based on cryo-electron microscopy maps and the interaction properties were estimated by means of a molecular dynamics approach, which simulate the unbinding of the complex applying a virtual spring to the core of myosin protein. Our results suggest that during an ATP hydrolysis cycle the affinity of myosin for actin is modulated by the presence and nature of the nucleotide in the active site of the myosin motor domain. When performing unbinding simulations with a pulling rate of 0.001 nm/ps, the maximum pulling force applied to the myosin during the experiment is about 1nN. Under these conditions the interaction force between myosin and actin monomer decreases from 0.83 nN in the nucleotide-free state to 0.27 nN in the ATP state, and increases to 0.60 nN after ATP hydrolysis and Pi release from the complex (ADP state)