Generation and modulation of network oscillations on the rodent prefrontal cortex in vitro

PhD ThesisFast network oscillations (~12-80 Hz) are recorded extensively in the mammalian cerebral cortex in vivo which local and distant neuronal populations orchestrate their firing activity to process cognitive-related information. The rat medial prefrontal cortex (mPFC) is considered to be functionally and anatomically homologous to the primate in vitro studies have demonstrated that the mPFC can sustain carbachol-induced persistent beta1 or kainate-induced transient low gamma frequency oscillations. We wished to establish an in vitro paradigm of carbachol (10 μM) / kainate (200 objective to investigate the distribution patterns and the mechanisms of these oscillations. Then we assessed the modulatory effects of the ascending catecholamine systems on fast network oscillations with exogenous application of Persistent fast network oscillations in the ventral mPFC were stronger, more rhythmic but slower (~25 Hz) than oscillations in the dorsal mPFC (~28 Hz). The regional difference in the oscillation amplitude was correlated to the strong regions in the mPFC, oscillations were stronger in layer 5. Oscillations relied on GABA, kainate but not AMPA receptors. In the ventral mPFC, network oscillations A were also dependent on NMDA receptor-mediated synaptic transmission. μM) reduced the oscillation strength and rhythmicity in the ventral mPFC. Instead, dopamine increased the power and rhythmicity of network oscillations in the dorsal mPFC. The region-dependent dopamine effect was correlated to the induced effects on synaptic inhibition and neuronal firing. μM) reduced the osc caused no effect on the dorsal mPFC

On the propensity of Asn-Gly-containing heptapeptides to form β\beta-turn structures : comparison between ab initio quantum mechanical calculations and Molecular Dynamics simulations

Both molecular mechanical and quantum mechanical calculations play an important role in describing the behavior and structure of molecules. In this work, we compare for the same peptide systems the results obtained from folding molecular dynamics simulations with previously reported results from quantum mechanical calculations. More specifically, three molecular dynamics simulations of 5 $\mu$s each in explicit water solvent were carried out for three Asn-Gly-containing heptapeptides, in order to study their folding and dynamics. Previous data, based on quantum mechanical calculations and the DFT methods have shown that these peptides adopt $\beta$-turn structures in aqueous solution, with type I' $\beta$-turn being the most preferred motif. The results from our analyses indicate that for the given system the two methods diverge in their predictions. The possibility of a force field-dependent deficiency is examined as a possible source of the observed discrepancy.Comment: In the supplementary information file figures S5, S6 and S7 the $\beta$-turn occupancies were wrong. They have been correcte

Designing a gamified social platform for people living with dementia and their live-in family caregivers

In the current paper, a social gamified platform for people living with dementia and their live-in family caregivers, integrating a broader diagnostic approach and interactive interventions is presented. The CAREGIVERSPRO-MMD (C-MMD) platform constitutes a support tool for the patient and the informal caregiver - also referred to as the dyad - that strengthens self-care, and builds community capacity and engagement at the point of care. The platform is implemented to improve social collaboration, adherence to treatment guidelines through gamification, recognition of progress indicators and measures to guide management of patients with dementia, and strategies and tools to improve treatment interventions and medication adherence. Moreover, particular attention was provided on guidelines, considerations and user requirements for the design of a User-Centered Design (UCD) platform. The design of the platform has been based on a deep understanding of users, tasks and contexts in order to improve platform usability, and provide adaptive and intuitive User Interfaces with high accessibility. In this paper, the architecture and services of the C-MMD platform are presented, and specifically the gamification aspects. © 2018 Association for Computing Machinery.Peer ReviewedPostprint (author's final draft

Structure of HrcQ(B)-C, a conserved component of the bacterial type III secretion systems

Type III secretion systems enable plant and animal bacterial pathogens to deliver virulence proteins into the cytosol of eukaryotic host cells, causing a broad spectrum of diseases including bacteremia, septicemia, typhoid fever, and bubonic plague in mammals, and localized lesions, systemic wilting, and blights in plants. In addition, type III secretion systems are also required for biogenesis of the bacterial flagellum. The HrcQ(B) protein, a component of the secretion apparatus of Pseudomonas syringae with homologues in all type III systems, has a variable N-terminal and a conserved C-terminal domain (HrcQ(B)-C). Here, we report the crystal structure of HrcQ(B)-C and show that this domain retains the ability of the full-length protein to interact with other type III components. A 3D analysis of sequence conservation patterns reveals two clusters of residues potentially involved in protein–protein interactions. Based on the analogies between HrcQ(B) and its flagellum homologues, we propose that HrcQ(B)-C participates in the formation of a C-ring-like assembly

Evolution of substrate specificity in a recipient's enzyme following horizontal gene transfer

Despite the prominent role of horizontal gene transfer (HGT) in shaping bacterial metabolism, little is known about the impact of HGT on the evolution of enzyme function. Specifically, what is the influence of a recently acquired gene on the function of an existing gene? For example, certain members of the genus Corynebacterium have horizontally acquired a whole L-tryptophan biosynthetic operon, whereas in certain closely related actinobacteria, for example, Mycobacterium, the trpF gene is missing. In Mycobacterium, the function of the trpF gene is performed by a dual-substrate (βα)8 phosphoribosyl isomerase (priA gene) also involved in L-histidine (hisA gene) biosynthesis. We investigated the effect of a HGT-acquired TrpF enzyme upon PriA’s substrate specificity in Corynebacterium through comparative genomics and phylogenetic reconstructions. After comprehensive in vivo and enzyme kinetic analyses of selected PriA homologs, a novel (βα)8 isomerase subfamily with a specialized function in L-histidine biosynthesis, termed subHisA, was confirmed. X-ray crystallography was used to reveal active-site mutations in subHisA important for narrowing of substrate specificity, which when mutated to the naturally occurring amino acid in PriA led to gain of function. Moreover, in silico molecular dynamic analyses demonstrated that the narrowing of substrate specificity of subHisA is concomitant with loss of ancestral protein conformational states. Our results show the importance of HGT in shaping enzyme evolution and metabolism

Order through Disorder: Hyper-Mobile C-Terminal Residues Stabilize the Folded State of a Helical Peptide. A Molecular Dynamics Study

Conventional wisdom has it that the presence of disordered regions in the three-dimensional structures of polypeptides not only does not contribute significantly to the thermodynamic stability of their folded state, but, on the contrary, that the presence of disorder leads to a decrease of the corresponding proteins' stability. We have performed extensive 3.4 µs long folding simulations (in explicit solvent and with full electrostatics) of an undecamer peptide of experimentally known helical structure, both with and without its disordered (four residue long) C-terminal tail. Our simulations clearly indicate that the presence of the apparently disordered (in structural terms) C-terminal tail, increases the thermodynamic stability of the peptide's folded (helical) state. These results show that at least for the case of relatively short peptides, the interplay between thermodynamic stability and the apparent structural stability can be rather subtle, with even disordered regions contributing significantly to the stability of the folded state. Our results have clear implications for the understanding of peptide energetics and the design of foldable peptides

The rotation-coupled sliding of EcoRV

It has been proposed that certain type II restriction enzymes (REs), such as EcoRV, track the helical pitch of DNA as they diffuse along DNA, a so-called rotation-coupled sliding. As of yet, there is no direct experimental observation of this phenomenon, but mounting indirect evidence gained from single-molecule imaging of RE–DNA complexes support the hypothesis. We address this issue by conjugating fluorescent labels of varying size (organic dyes, proteins and quantum dots) to EcoRV, and by fusing it to the engineered Rop protein scRM6. Single-molecule imaging of these modified EcoRVs sliding along DNA provides us with their linear diffusion constant (D1), revealing a significant size dependency. To account for the dependence of D1 on the size of the EcoRV label, we have developed four theoretical models describing different types of motion along DNA and find that our experimental results are best described by rotation-coupled sliding of the protein. The similarity of EcoRV to other type II REs and DNA binding proteins suggests that this type of motion could be widely preserved in other biological contexts