The induction of α-helical structure in partially unfolded HypF-N does not affect its aggregation propensity

The conversion of proteins into structured fibrillar aggregates is a central problem in protein chemistry, biotechnology, biology and medicine. It is generally accepted that aggregation takes place from partially structured states of proteins. However, the role of the residual structure present in such conformational states is not yet understood. In particular, it is not yet clear as to whether the α-helical structure represents a productive or counteracting structural element for protein aggregation. We have addressed this issue by studying the aggregation of pH-unfolded HypF-N. It has previously been shown that the two native α-helices of HypF-N retain a partial α-helical structure in the pH-unfolded state and that these regions are also involved in the formation of the cross-β structure of the aggregates. We have introduced mutations in such stretches of the sequence, with the aim of increasing the α-helical structure in the key regions of the pH-unfolded state, while minimizing the changes of other factors known to influence protein aggregation, such as hydrophobicity, β-Sheet propensity, etc. The resulting HypF-N mutants have higher contents of α-helical structure at the site(s) of mutation in their pH-unfolded states, but such an increase does not correlate with a change of aggregation rate. The results suggest that stabilisation of α-helical structure in amyloidogenic regions of the sequence of highly dynamic states does not have remarkable effects on the rate of protein aggregation from such conformational states. Comparison with other protein systems indicate that the effect of increasing α-helical propensity can vary if the stabilised helices are in non-amyloidogenic stretches of initially unstructured peptides (accelerating effect), in amyloidogenic stretches of initially unstructured peptides (no effect) or in amyloidogenic stretches of initially stable helices (decelerating effect

BoBafit: A copy number clustering tool designed to refit and recalibrate the baseline region of tumors’ profiles

Human cancer arises from a population of cells that have acquired a wide range of genetic alterations, most of which are targets of therapeutic treatments or are used as prognostic factors for patient's risk stratification. Among these, copy number alterations (CNAs) are quite frequent. Currently, several molecular biology technologies, such as microarrays, NGS and single-cell approaches are used to define the genomic profile of tumor samples. Output data need to be analyzed with bioinformatic approaches and particularly by employing computational algorithms. Molecular biology tools estimate the baseline region by comparing either the mean probe signals, or the number of reads to the reference genome. However, when tumors display complex karyotypes, this type of approach could fail the baseline region estimation and consequently cause errors in the CNAs call. To overcome this issue, we designed an R-package, BoBafit, able to check and, eventually, to adjust the baseline region, according to both the tumor-specific alterations’ context and the sample-specific clustered genomic lesions. Several databases have been chosen to set up and validate the designed package, thus demonstrating the potential of BoBafit to adjust copy number (CN) data from different tumors and analysis techniques. Relevantly, the analysis highlighted that up to 25% of samples need a baseline region adjustment and a redefinition of CNAs calls, thus causing a change in the prognostic risk classification of the patients. We support the implementation of BoBafit within CN analysis bioinformatics pipelines to ensure a correct patient's stratification in risk categories, regardless of the tumor type

Microbial Community Characterizing Vermiculations from Karst Caves and Its Role in Their Formation

The microbiota associated with vermiculations from karst caves is largely unknown. Vermiculations are enigmatic deposits forming worm-like patterns on cave walls all over the world. They represent a precious focus for geomicrobiological studies aimed at exploring both the microbial life of these ecosystems and the vermiculation genesis. This study comprises the first approach on the microbial communities thriving in Pertosa-Auletta Cave (southern Italy) vermiculations by next-generation sequencing. The most abundant phylum in vermiculations was Proteobacteria, followed by Acidobacteria > Actinobacteria > Nitrospirae > Firmicutes > Planctomycetes > Chloroflexi > Gemmatimonadetes > Bacteroidetes > Latescibacteria. Numerous less-represented taxonomic groups (< 1%), as well as unclassified ones, were also detected. From an ecological point of view, all the groups co-participate in the biogeochemical cycles in these underground environments, mediating oxidation-reduction reactions, promoting host rock dissolution and secondary mineral precipitation, and enriching the matrix in organic matter. Confocal laser scanning microscopy and field emission scanning electron microscopy brought evidence of a strong interaction between the biotic community and the abiotic matrix, supporting the role of microbial communities in the formation process of vermiculations

Identification of a Maturation Plasma Cell Index through a Highly Sensitive Droplet Digital PCR Assay Gene Expression Signature Validation in Newly Diagnosed Multiple Myeloma Patients

DNA microarrays and RNA-based sequencing approaches are considered important discovery tools in clinical medicine. However, cross-platform reproducibility studies undertaken so far have highlighted that microarrays are not able to accurately measure gene expression, particularly when they are expressed at low levels. Here, we consider the employment of a digital PCR assay (ddPCR) to validate a gene signature previously identified by gene expression profile. This signature included ten Hedgehog (HH) pathways' genes able to stratify multiple myeloma (MM) patients according to their self-renewal status. Results show that the designed assay is able to validate gene expression data, both in a retrospective as well as in a prospective cohort. In addition, the plasma cells' differentiation status determined by ddPCR was further confirmed by other techniques, such as flow cytometry, allowing the identification of patients with immature plasma cells' phenotype (i.e., expressing CD19+/CD81+ markers) upregulating HH genes, as compared to others, whose plasma cells lose the expression of these markers and were more differentiated. To our knowledge, this is the first technical report of gene expression data validation by ddPCR instead of classical qPCR. This approach permitted the identification of a Maturation Index through the integration of molecular and phenotypic data, able to possibly define upfront the differentiation status of MM patients that would be clinically relevant in the future

Antibacterial Al-doped ZnO coatings on PLA films

Antimicrobial surfaces can play a key role in many fields, like biomedical applications and food packaging, where it is fundamental to prevent clinical infections, foodborne diseases, or quality loss of food and beverages. Materials used for such kind of applications should be biocompatible or avoid harmful effects on human health. Additionally, since a wide variety of tools and components operating in contact with human tissues or foodstuff are made of plastic-based materials, it is important to investigate antibacterial surfaces applied in conjunction with eco-friendly bio-based plastics. For all these reasons, here we propose the use of novel composites made of nanostructured aluminum-doped zinc oxide antimicrobial coatings on polylactide films. Their surface topography is inspected at sub- and supra-micron length scales, and its influence on wettability and antibacterial activity is investigated, together with FTIR analysis for identification of molecular vibrations. Their strong antimicrobial performance is assessed against four different bacterial strains, and their safety is evaluated on two different human cell lines. The results here presented identify AZO-coated PLA films as very appealing eco-friendly antibacterial systems for food packaging and biomedical applications

New compounds for a good old class: Synthesis of two Β-lactam bearing cephalosporins and their evaluation with a multidisciplinary approach

Antimicrobial resistance is spreading massively in the world and is becoming one of the main health threats of the 21st century. One of the possible strategies to overcome this problem is to modify the known classes of antibiotics in a rational way, with the aim of tuning their efficacy. In this paper, we present the synthesis and the evaluation of the biological activity of a series of two β-lactam bearing cephalosporin derivatives, in which an additional isolated azetidinone ring, bearing different substituents, is joined to the classical cephalosporanic nucleus by a chain of variable length. A computational approach has been also applied in order to predict the molecular interactions between some representative derivatives and selected penicillin-binding proteins, the natural targets of β-lactam antibiotics. All these derivatives are active against Gram-positive bacteria, with MIC100 comparable or even better than that of the reference antibiotic ceftriaxone, and show no or very low cytotoxic activity on different cell lines. Overall, these molecules appear to be able to exert their activity in particular against microorganisms belonging to some of the species more involved in the development of multidrug resistance