Modulation of the stability of the Salmonella fourU-type RNA thermometer

RNA thermometers are translational control elements that regulate the expression of bacterial heat shock and virulence genes. They fold into complex secondary structures that block translation at low temperatures. A temperature increase releases the ribosome binding site and thus permits translation initiation. In fourU-type RNA thermometers, the AGGA sequence of the SD region is paired with four consecutive uridines. We investigated the melting points of the wild-type and mutant sequences. It was decreased by 5°C when a stabilizing GC basepair was exchanged by an AU pair or increased by 11°C when an internal AG mismatch was converted to a GC pair, respectively. Stabilized or destabilized RNA structures are directly correlated with decreased or increased in vivo gene expression, respectively. Mg2+ also affected the melting point of the fourU thermometer. Variations of the Mg2+ concentration in the physiological range between 1 and 2 mM translated into a 2.8°C shift of the melting point. Thus, Mg2+ binding to the hairpin RNA is regulatory relevant. Applying three different NMR techniques, two Mg2+ binding sites were found in the hairpin structure. One of these binding sites could be identified as outer sphere binding site that is located within the fourU motif. Binding of the two Mg2+ ions exhibits a positive cooperativity with a Hill coefficient of 1.47. Free energy values delta G for Mg2+ binding determined by NMR are in agreement with data determined from CD measurements. Physiological Mg2+ concentrations reduce enthalpy and entropy values of uncorrelated base pair opening processes for almost all nucleobases

Exploration of the scalability of LocFaults approach for error localization with While-loops programs

A model checker can produce a trace of counterexample, for an erroneous program, which is often long and difficult to understand. In general, the part about the loops is the largest among the instructions in this trace. This makes the location of errors in loops critical, to analyze errors in the overall program. In this paper, we explore the scala-bility capabilities of LocFaults, our error localization approach exploiting paths of CFG(Control Flow Graph) from a counterexample to calculate the MCDs (Minimal Correction Deviations), and MCSs (Minimal Correction Subsets) from each found MCD. We present the times of our approach on programs with While-loops unfolded b times, and a number of deviated conditions ranging from 0 to n. Our preliminary results show that the times of our approach, constraint-based and flow-driven, are better compared to BugAssist which is based on SAT and transforms the entire program to a Boolean formula, and further the information provided by LocFaults is more expressive for the user

Constraint-Based Qualitative Simulation

We consider qualitative simulation involving a finite set of qualitative relations in presence of complete knowledge about their interrelationship. We show how it can be naturally captured by means of constraints expressed in temporal logic and constraint satisfaction problems. The constraints relate at each stage the 'past' of a simulation with its 'future'. The benefit of this approach is that it readily leads to an implementation based on constraint technology that can be used to generate simulations and to answer queries about them.Comment: 10 pages, to appear at the conference TIME 200

A synchronous program algebra: a basis for reasoning about shared-memory and event-based concurrency

This research started with an algebra for reasoning about rely/guarantee concurrency for a shared memory model. The approach taken led to a more abstract algebra of atomic steps, in which atomic steps synchronise (rather than interleave) when composed in parallel. The algebra of rely/guarantee concurrency then becomes an instantiation of the more abstract algebra. Many of the core properties needed for rely/guarantee reasoning can be shown to hold in the abstract algebra where their proofs are simpler and hence allow a higher degree of automation. The algebra has been encoded in Isabelle/HOL to provide a basis for tool support for program verification. In rely/guarantee concurrency, programs are specified to guarantee certain behaviours until assumptions about the behaviour of their environment are violated. When assumptions are violated, program behaviour is unconstrained (aborting), and guarantees need no longer hold. To support these guarantees a second synchronous operator, weak conjunction, was introduced: both processes in a weak conjunction must agree to take each atomic step, unless one aborts in which case the whole aborts. In developing the laws for parallel and weak conjunction we found many properties were shared by the operators and that the proofs of many laws were essentially the same. This insight led to the idea of generalising synchronisation to an abstract operator with only the axioms that are shared by the parallel and weak conjunction operator, so that those two operators can be viewed as instantiations of the abstract synchronisation operator. The main differences between parallel and weak conjunction are how they combine individual atomic steps; that is left open in the axioms for the abstract operator.Comment: Extended version of a Formal Methods 2016 paper, "An algebra of synchronous atomic steps

A synchronous program algebra: a basis for reasoning about shared-memory and event-based concurrency

This research started with an algebra for reasoning about rely/guarantee concurrency for a shared memory model. The approach taken led to a more abstract algebra of atomic steps, in which atomic steps synchronise (rather than interleave) when composed in parallel. The algebra of rely/guarantee concurrency then becomes an instantiation of the more abstract algebra. Many of the core properties needed for rely/guarantee reasoning can be shown to hold in the abstract algebra where their proofs are simpler and hence allow a higher degree of automation. The algebra has been encoded in Isabelle/HOL to provide a basis for tool support for program verification. In rely/guarantee concurrency, programs are specified to guarantee certain behaviours until assumptions about the behaviour of their environment are violated. When assumptions are violated, program behaviour is unconstrained (aborting), and guarantees need no longer hold. To support these guarantees a second synchronous operator, weak conjunction, was introduced: both processes in a weak conjunction must agree to take each atomic step, unless one aborts in which case the whole aborts. In developing the laws for parallel and weak conjunction we found many properties were shared by the operators and that the proofs of many laws were essentially the same. This insight led to the idea of generalising synchronisation to an abstract operator with only the axioms that are shared by the parallel and weak conjunction operator, so that those two operators can be viewed as instantiations of the abstract synchronisation operator. The main differences between parallel and weak conjunction are how they combine individual atomic steps; that is left open in the axioms for the abstract operator.Comment: Extended version of a Formal Methods 2016 paper, "An algebra of synchronous atomic steps

Constraint Design Rewriting

We propose an algebraic approach to the design and transformation of constraint networks, inspired by Architectural Design Rewriting. The approach can be understood as (i) an extension of ADR with constraints, and (ii) an application of ADR to the design of reconfigurable constraint networks. The main idea is to consider classes of constraint networks as algebras whose operators are used to denote constraint networks with terms. Constraint network transformations such as constraint propagations are specified with rewrite rules exploiting the network’s structure provided by terms

Structural and functional characterization of a cold-adapted stand-alone TPM domain reveals a relationship between dynamics and phosphatase activity

The TPM domain constitutes a family of recently characterized protein domains that are present in most living organisms. Although some progress has been made in understanding the cellular role of TPM-containing proteins, the relationship between structure and function is not clear yet. We have recently solved the solution and crystal structure of one TPM domain (BA42) from the Antarctic bacterium Bizionia argentinensis. In this work, we demonstrate that BA42 has phosphoric-monoester hydrolase activity. The activity of BA42 is strictly dependent on the binding of divalent metals and retains nearly 70% of the maximum at 4 °C, a typical characteristic of cold-adapted enzymes. From HSQC, 15 N relaxation measurements, and molecular dynamics studies, we determine that the flexibility of the crossing loops was associated to the protein activity. Thermal unfolding experiments showed that the local increment in flexibility of Mg2+ -bound BA42, when compared with Ca2+ -bound BA42, is associated to a decrease in global protein stability. Finally, through mutagenesis experiments, we unambiguously demonstrate that the region comprising the metal-binding site participates in the catalytic mechanism. The results shown here contribute to the understanding of the relationship between structure and function of this new family of TPM domains providing important cues on the regulatory role of Mg2+ and Ca2+ and the molecular mechanism underlying enzyme activity at low temperatures.Fil: Pellizza, Leonardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Smal, Clara. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Ithuralde, Raúl Esteban. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales; ArgentinaFil: Turjanski, Adrian. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales; ArgentinaFil: Cicero, Daniel Oscar. Universita Tor Vergata; Italia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Aran, Martin. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; Argentin

A general theory of action languages

We present a general theory of action-based languages as a paradigm, for the description, of those computational systems which include elements of concurrency and networking, and extend this approach to describe dist.ributed systems and also t,o describe the interaction of a system, with an environment. As part of this approach we introduce the Action Language as a common model for the class of nondeterministic concurrent programming languages and define its intensional and interaction semantics in terrors of continuous transformation of environment behavior. This semantics i.s specialized for programs with stores, and extended to describe distributed computations