Necessity is the Mother of Invention: A Remote Molecular Bioinformatics Practical Course in the COVID-19 Era.

The COVID-19 pandemic has brought many challenges to human beings, related to not only health and way of life but also teaching because of the interruption of the standard training at universities imposed by lockdowns. Concerning the latter, the academic community had to reinvent itself, in many ways, to carry on with prepandemic education. This article focuses on the use of modern technology and software to create a virtual, highly interactive classroom where a remote but still hands-on course on molecular bioinformatics can be taught, motivating the university students and helping them learn the course contents without significant compromises imposed by successive lockdowns. We implemented such a virtual hands-on molecular bioinformatics course in the second semester of the 2020/2021 academic year. Furthermore, we compared the learning outcomes with those for the earlier editions of the same course in the pre-COVID-19 era, in which the more traditional teaching method was used where all teaching was delivered with physically present lecturers. The virtual classroom proposed here allowed the students to develop skills close to, although slightly below, those obtained with physically present learning.

Lessons from a Single Amino Acid Substitution: Anticancer and Antibacterial Properties of Two Phospholipase A2-Derived Peptides.

The membrane-active nature of phospholipase A2-derived peptides makes them potential candidates for antineoplastic and antibacterial therapies. Two short 13-mer C-terminal fragments taken from snake venom Lys49-PLA2 toxins (p-AppK and p-Acl), differing by a leucine/phenylalanine substitution, were synthesized and their bioactivity was evaluated. Their capacity to interfere with the survival of Gram-positive and Gram-negative bacteria as well as with solid and liquid tumors was assessed in vitro. Toxicity to red blood cells was investigated via in silico and in vitro techniques. The mode of action was mainly studied by molecular dynamics simulations and membrane permeabilization assays. Briefly, both peptides have dual activity, i.e., they act against both bacteria, including multidrug-resistant strains and tumor cells. All tested bacteria were susceptible to both peptides, Pseudomonas aeruginosa being the most affected. RAMOS, K562, NB4, and CEM cells were the main leukemic targets of the peptides. In general, p-Acl showed more significant activity, suggesting that phenylalanine confers advantages to the antibacterial and antitumor mechanism, particularly for osteosarcoma lines (HOS and MG63). Peptide-based treatment increased the uptake of a DNA-intercalating dye by bacteria, suggesting membrane damage. Indeed, p-AppK and p-Acl did not disrupt erythrocyte membranes, in agreement with in silico predictions. The latter revealed that the peptides deform the membrane and increase its permeability by facilitating solvent penetration. This phenomenon is expected to catalyze the permeation of solutes that otherwise could not cross the hydrophobic membrane core. In conclusion, the present study highlights the role of a single amino acid substitution present in natural sequences towards the development of dual-action agents. In other words, dissecting and fine-tuning biomembrane remodeling proteins, such as snake venom phospholipase A2 isoforms, is again demonstrated as a valuable source of therapeutic peptides.

A novel synthetic peptide inspired on Lys49 phospholipase A2 from Crotalus oreganus abyssus snake venom active against multidrug-resistant clinical isolates.

Currently, the evolving and complex mechanisms of bacterial resistance to conventional antibiotics are increasing, while alternative medicines are drying up, which urges the need to discover novel agents able to kill antibiotic-resistant bacteria. Lys49 phospholipase A2s (PLA2s) from snake venoms are multifunctional toxins able to induce a huge variety of therapeutic effects and consequently serve as templates for new drug leads. Hence, the present study was aimed at the synthesis of oligopeptides mimicking regions of the antibacterial Lys49 PLA2 toxin (CoaTx-II), recently isolated from Crotalus oreganus abyssus snake venom, to identify small peptides able to reproduce the therapeutic action of the toxin. Five peptides, representing major regions of interest within CoaTx-II, were synthesized and screened for their antibacterial properties. The 13-mer peptide pC-CoaTxII, corresponding to residues 115-129 of CoaTx-II, was able to reproduce the promising bactericidal effect of the toxin against multi-resistant clinical isolates. Peptide pC-CoaTxII is mainly composed by positively charged and hydrophobic amino acids, a typical trait in most antimicrobial peptides, and presented no defined secondary structure in aqueous environment. The physicochemical properties of pC-CoaTxII are favorable towards a strong interaction with anionic lipid membranes as those in bacteria. Additional in silico studies suggest formation of a water channel across the membrane upon peptide insertion, eventually leading to bacterial cell disruption and death. Overall, our findings confirm the valuable potential of snake venom toxins towards design and synthesis of novel antimicrobials, thus representing key insights towards development of alternative efficient antimicrobials to fight bacterial resistance to current antibiotics.

QM/MM study of the catalytic mechanism of GalNAc removal from GM2 ganglioside catalyzed by human ß-hexosaminidaseA.

This work is based on the glycosidase ß-hexosaminidase, in particular, on ß-HexA (HexA), involved in the emergence of the Sandoff disease. Its function is to cleave the N-acetylgalactosamine (GalNAc) or N-acetylglucosamine moieties from its substrates. Here we reveal and consolidate many important aspects of the catalytic mechanism of GalNAc removal from the GM2 ganglioside. The reaction mechanism for this reaction is proposed to be substrate-assisted. It is expected to evolve through a mechanism similar to others glycosidase mechanisms, even though this fact has never been confirmed and the proposal still lacks atomic level detail in its description. To overcome these two limitations we have used the ONIOM formalism (B3LYP:Amber and M06-2X:Amber) with electrostatic embedding to calculate a bidimensional potential energy surface for the rate-limiting step. The potential energy surface reveals the mechanism with atomic detail. The formation of the covalent bond within the substrate has been confirmed, and its energy of stabilization has been calculated as 5.1 kcal/mol. The electronic energy barrier for this step is 22.5 kcal/mol at the hybrid ONIOM(M06-2X:Amber) level, well in line with the typical experimental values for similar reactions in glycosylases. The conformational transitions of the substrate GalNAc ring have been mapped in the PES for the first time. They support the hypothesis of a (4)S(2) skew boat-(4)E envelope-(4)C(1) chair pathway and provide much finer detail to the earlier proposals. In general the results are in line with earlier predictions.

Doença de Still no adulto: relato de um caso / Still's disease in the adult: report of a case

Os autores apresentam um caso de Doença de Still no adulto, acometendo homem de 35 anos de idade, com diagnóstico prévio de Febre Reumática. Fazem uma abordagem terapêutica clássica e um estudo bibliográfico sobre o tema