Antonio Caliri

Phospholipase A2 Isolated from the Venom of Crotalus durissus terrificus Inactivates Dengue virus and Other Enveloped Viruses by Disrupting the Viral Envelope

The Flaviviridae family includes several virus pathogens associated with human diseases worldwide. Within this family, Dengue virus is the most serious threat to public health, especially in tropical and sub-tropical regions of the world. Currently, there are no vaccines or specific antiviral drugs against Dengue virus or against most of the viruses of this family. Therefore, the development of vaccines and the discovery of therapeutic compounds against the medically most important flaviviruses remain a global public health priority. We previously showed that phospholipase A2 isolated from the venom of Crotalus durissus terrificus was able to inhibit Dengue virus and Yellow fever virus infection in Vero cells. Here, we present evidence that phospholipase A2 has a direct effect on Dengue virus particles, inducing a partial exposure of genomic RNA, which strongly suggests inhibition via the cleavage of glycerophospholipids at the virus lipid bilayer envelope. This cleavage might induce a disruption of the lipid bilayer that causes a destabilization of the E proteins on the virus surface, resulting in inactivation. We show by computational analysis that phospholipase A2 might gain access to the Dengue virus lipid bilayer through the pores found on each of the twenty 3-fold vertices of the E protein shell on the virus surface. In addition, phospholipase A2 is able to inactivate other enveloped viruses, highlighting its potential as a natural product lead for developing broad-spectrum antiviral drugs.

Deterministic folding: The role of entropic forces and steric specificities

The inverse folding problem of proteinlike macromolecules is studied by using a lattice Monte Carlo (MC) model in which steric specificities (nearest-neighbors constraints) are included and the hydrophobic effect is treated explicitly by considering interactions between the chain and solvent molecules. Chemical attributes and steric peculiarities of the residues are encoded in a 10-letter alphabet and a correspondent “syntax” is provided in order to write suitable sequences for the specified target structures; twenty-four target configurations, chosen in order to cover all possible values of the average contact order χ (0.2381⩽χ⩽0.4947 for this system), were encoded and analyzed. The results, obtained by MC simulations, are strongly influenced by geometrical properties of the native configuration, namely χ and the relative number φ of crankshafts-type structures: For χ<0.35 the folding is deterministic, that is, the syntax is able to encode successful sequences: The system presents larger encodability, mi...

Nonextensive statistical mechanics applied to protein folding problem: kinetics aspects

A reduced (stereo-chemical) model is employed to study kinetic aspects of globular protein folding process, by Monte Carlo simulation. Nonextensive statistical approach is used: transition probability pi j between configurations i → j is given by pi j =[1 +(1 - q)ΔGi j/kBT ]1/(1-q), where q is the nonextensive (Tsallis) parameter. The system model consists of a chain of 27 beads immerse in its solvent; the beads represent the sequence of amino acids along the chain by means of a 10-letter stereo-chemical alphabet; a syntax (rule) to design the amino acid sequence for any given 3D structure is embedded in the model. The study focuses mainly kinetic aspects of the folding problem related with the protein folding time, represented in this work by the concept of first passage time (FPT). Many distinct proteins, whose native structures are represented here by compact self avoiding (CSA) configurations, were employed in our analysis, although our results are presented exclusively for one representative protein, for which a rich statistics was achieved. Our results reveal that there is a specific combinations of value for the nonextensive parameter q and temperature T, which gives the smallest estimated folding characteristic time (t). Additionally, for q = 1.1, (t) stays almost invariable in the range 0.9 < T < 1.3, slightly oscillating about its average value = 27 ±σ, where σ = 2 is the standard deviation. This behavior is explained by comparing the distribution of the folding times for the Boltzmann statistics (q → 1), with respect to the nonextensive statistics for q = 1.1, which shows that the effect of the nonextensive parameter q is to cut off the larger folding times present in the original (q → 1) distribution. The distribution of natural logarithm of the folding times for Boltzmann statistics is a triple peaked Gaussian, while, for q = 1.1 (Tsallis), it is a double peaked Gaussian, suggesting that a log-normal process with two characteristic times replaced the original process with three characteristic times. Finally we comment on the physical meaning of the present results, as well its significance in the near future works.

Membrane vesiculation induced by proteins of the dengue virus envelope studied by molecular dynamics simulations

Abstract Biological membranes are continuously remodeled in the cell by specific membrane-shaping machineries to form, for example, tubes and vesicles. We examine fundamental mechanisms involved in the vesiculation processes induced by a cluster of envelope (E) and membrane (M) proteins of the dengue virus (DENV) using molecular dynamics simulations and a coarse-grained model. We show that an arrangement of three E-M heterotetramers (EM3) works as a bending unit and an ordered cluster of five such units generates a closed vesicle, reminiscent of the virus budding process. In silico mutagenesis of two charged residues of the anchor helices of the envelope proteins of DENV shows that Arg-471 and Arg-60 are fundamental to produce bending stress on the membrane. The fine-tuning between the size of the EM3 unit and its specific bending action suggests this protein unit is an important factor in determining the viral particle size.

Baccharis dracunculifolia DC (Asteraceae) selectively modulates the effector functions of human neutrophils

To examine whether the hydroalcoholic extract from Baccharis dracunculifolia leaves (BdE) modulates the human neutrophil oxidative metabolism, degranulation, phagocytosis and microbial killing capacity.

In silico assessment of S100A12 monomer and dimer structural dynamics: implications for the understanding of its metal-induced conformational changes

Changes in the concentration of different ions modulate several cellular processes, such as Ca2+ and Zn2+ in inflammation. Upon activation of immune system effector cells, the intracellular Ca2+ concentration rises propagating the activation signal, leading to degranulation and generation of reactive oxygen species, which increases the Zn2+ intracellular concentration as a consequence of the cellular antioxidant machinery. In this context, S100A12 is of special interest because it is a pro-inflammatory protein expressed in neutrophils whose structure and function are modulated by both Ca2+ and Zn2+. The current hypothesis about its mechanism of action was built based on biochemical and crystallographic data. However, there are missing connections between molecular structure and the way in which many events are concatenated at the triggering and along the inflammatory process. In this work we use molecular dynamics simulations to describe how variations in Zn2+ and Ca2+ concentrations modulate the structural dynamics of the calcium-free S100A12 dimer and monomer, which was not considered a part of the mechanism of action before. Our results suggest that (i) Zn2+ have a determinant role in the dimerization step, as well as in the unbinding of the Na+ complexed to the N-terminal EF-hand; (ii) the N-terminal EF-hand domain is the first to bind Ca2+, and not the C-terminal, as usually accepted; and that (iii) Ca2+ modulates the structural dynamics of H-III.

The 3-phenylcoumarin derivative 6,7-dihydroxy-3-[3′,4′-methylenedioxyphenyl]-coumarin downmodulates the FcγR- and CR-mediated oxidative metabolism and elastase release in human neutrophils: Possible mechanisms underlying inhibition of the formation and release of neutrophil extracellular traps

ABSTRACT In this study, we report the ability of a set of eight 3‐phenylcoumarin derivatives bearing 6,7‐ or 5,7‐dihydroxyl groups, free or acetylated, bound to the benzopyrone moiety, to modulate the effector functions of human neutrophils. In general, (i) 6,7‐disubstituted compounds (5, 6, 19, 20) downmodulated the Fc&ggr; receptor‐mediated neutrophil oxidative metabolism more strongly than 5,7‐disubstituted compounds (21, 22, 23, 24), and (ii) hydroxylated compounds (5, 19, 21, 23) downmodulated this neutrophil function more effectively than their acetylated counterparts (6, 20, 22, 24, respectively). Compounds 5 (6,7‐dihydroxy‐3‐[3′,4′‐methylenedioxyphenyl]‐coumarin) and 19 (6,7‐dihydroxy‐3‐[3′,4′‐dihydroxyphenyl]‐coumarin) effectively downmodulated the neutrophil oxidative metabolism elicited via Fc&ggr; and/or complement receptors. Compound 5 also downmodulated the immune complex‐stimulated phagocytosis, degranulation of elastase, and production and release of neutrophil extracellular traps, as well as the human neutrophil chemotaxis towards n‐formyl‐methionyl‐leucyl‐phenylalanine, without altering the expression level of formyl peptide receptor type 1. Both compounds 5 and 19 did not impair the neutrophil capacity to recognize and kill Candida albicans. Docking calculations revealed that compounds 5 and 19 directly interacted with three catalytic residues – Gln‐91, His‐95, and Arg‐239 – inside the myeloperoxidase active site. Together, these findings indicate that (i) inhibition of reactive oxygen species generation and degranulation of elastase are closely associated with downmodulation of release of neutrophil extracellular traps; and (ii) compound 5 can be a prototype for the development of novel immunomodulating drugs to treat immune complex‐mediated inflammatory diseases. Graphical abstract Figure. No caption available. Highlights3‐PD 5 downmodulates neutrophil ROS generation, phagocytosis, and degranulation.3‐PD 5 downmodulates the production and release of neutrophil extracellular traps.3‐PD 5 downmodulates neutrophil chemotaxis towards fMLP but not FPR1 expression.3‐PD 5 is not cytotoxic to neutrophils and does not interfere with microbial killing.3‐PD 5 directly interacts with three catalytic residues of myeloperoxidase.

Thermal properties of the ±J infinite-range Sherrington-Kirkpatrick model by analysis of the partition function zeros

Abstract An explicit and simple (approximated) solution of the ± J infinite-range Sherrington-Kirkpatrick (SK) model of spin glasses is presented: standard thermodynamic functions, valid for all temperatures, are derived by analysis of the partition function zeros in the complex temperature plane. Learning from computer calculations in finite systems, general thermodynamic considerations and a subsidiary heuristic assumption are used. The ground state energy is found to be −(6 log 2+5)/12 ≈ −0.76324, and the low-temperature specific heat as well the entropy are quadratic: 1.5 T 2 and 0.75 T 2 respectively.