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Vegetable wastes represent an inexpensive and sustainable source of valuable bioproducts for several applications. Natural micro-porous and fibrous materials can be obtained from a very cheap and abundant cellulosic bio-waste. Here we demonstrated that vegetable waste derivatives can be suitable as scaffolds for biosensors and 3D cell growth. Many studies have been addressed to fabricate biocompatible 3D scaffolds for mammalian stem cells cultures and develop novel systems able to reproduce the complexity of the in vivo microenvironment. Many of these products are proprietary, expensive or require chemical synthesis. The recycling and revaluation of vegetable derived tissues to fabricate scaffolds for analytical biosensors 3D stem cell cultures platforms may represent a very low-cost approach for toxicological and environmental analyses. In this approach, potential applications of vegetable-derived tissue for biosensing and 3D stem cell cultures were investigated. Micro-structured scaffolds from stalk of broccoli, named BrcS, were either functionalized for production of enzymatic 3D-biosensors or preconditioned to be used them as 3D-scaffolds for human mesenchymal stem cells cultures. The conditions to fabricate 3D-biosensors and scaffolds for cell growth were here optimized studying all analytical parameters and demonstrating the feasibility to combine these two properties for an innovative solution to ennoble vegetable wastes., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Hydrogels are good candidate materials for cell delivery scaffolds because they can mimic the physical, chemical, electrical and biological properties of most of the native tissues. In this study, composite biosynthetic hydrogels were produced by combining the bio-functionality of silk fibroin (SF) with the structural versatility of polyethylene-glycol-diacrylated (PEGDa). The formation of a photopolymerizable PEGDa-SF hydrogel (PSFHy) was optimized for 3D-cell culture. Functionalization of the 3D-PSFHy with protein microspheres (MS) was required to increase the porosity and cell-adhesive properties of the material. Cardiac mesenchymal stem cells, which were cultured within the MS-embedding PSFHy, exhibited good viability and expression of proteins that are characteristic of the initial phases of the cardiac muscle differentiation process. Further, the addition of chondroitin sulfate into the scaffolds improved the cell viability. A cell-preconditioning of the scaffold was also performed, suggesting a potential application of these sponge-like scaffolds for analysing the effects of several extracellular microenvironments, produced by different kinds of cells, on the stem cells fate. The results presented herein highlight on the possibility to use the PSFHys functionalized with MS as stem cell-carrier systems with sponge-like properties, potential ultrasound-imaging contrast agents and controlled biochemical factor delivery., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Background: An alarming rate of early failure has been recently reported for the LivaNova (previously Sorin) Mitroflow (LivaNova, London, UK) bioprosthesis. Here, we aimed at verifying if this possible underperformance is confirmed in a large, single-center experience and identifying the risk factors associated with early deterioration., Methods: In all, 459 Mitroflow valves have been implanted from July 2009 to December 2013 (patients' mean age 73 years; 204 women). Surviving patients have undergone yearly clinic and echocardiographic follow-up. Dysfunction was defined as moderate if the mean gradient was more than 30 mm Hg or severe if it exceeded 40 mm Hg. The population was divided on the basis of a dimensional mismatch, the model of the prosthesis (LX or DL: follow-up to 4 years), and patient's age at the time of implantation., Results: Cumulative freedom from moderate valve dysfunction was 81% ± 3% at 60 months. It was lower with patient-prosthesis mismatch (71% ± 5% versus 92% ± 3%; p = 0.0065) and with the more recent DL model (at 42 months: 78% ± 6% versus 96% ± 2%; p < 0.0001). Cumulative freedom from severe dysfunction was 93% ± 2% at 5 years. Again, it was inferior among patients with a mismatch (86% ± 4% versus 100%; p = 0.0013) and for the DL model (42 months: 92.5% ± 3% versus 98.5% ± 1%; p = 0.0309). Smaller prostheses showed higher rates of early degeneration., Conclusions: The LivaNova Mitroflow valve appears to be prone to early deterioration. Smaller size prostheses should be used cautiously and avoided with patient-prosthesis mismatch. The DL model anticalcification treatment seems unable to prevent early degeneration, and possibly contributes to even earlier failure., (Copyright © 2018 The Society of Thoracic Surgeons. Published by Elsevier Inc. All rights reserved.)

p63, a member of the p53 family, is a crucial transcription factor for epithelial development and skin homeostasis. Heterozygous mutations in TP63 gene have been associated with human ectodermal dysplasia disorders. Most of these TP63 mutations are missense mutations causing amino acidic substitutions at p63 DNA binding or SAM domains that reduce or abolish the transcriptional activity of mutants p63. A significant number of mutants, however, resides in part of the p63 protein that apparently do not affect DNA binding and/or transcriptional activity, such as the N-terminal domain. Here, we characterize five p63 mutations at the 5' end of TP63 gene aiming to understand the pathogenesis of the diseases and to uncover the role of ΔNp63α N-terminus residues in determining its transactivation potential., (Copyright © 2015 Elsevier Inc. All rights reserved.)

The human immunodeficiency virus type 1 aspartyl protease (HIV-1 PR) is a homodimeric aspartyl endopeptidase that is required for virus replication. HIV-1 PR was shown to act invitro as acyl-donor and -acceptor for both guinea pig liver transglutaminase (TG, EC 2.3.2.13) and human Factor XIIIa. These preliminary evidences suggested that the HIV-1 PR contains at least three TG-reactive glutaminyl and one lysyl residues. We report here that the incubation of HIV-1 PR with TG increases its catalytic activity. This increase is dependent upon the time of incubation, the concentration of TG and the presence of Ca2+. Identification of epsilon-(gamma-glutamyl)lysine in the proteolytic digest of the TG-modified HIV-1 PR suggested intramolecular covalent cross-linking of this protease which may promote a non-covalent dimerization and subsequent activation of this enzyme via a conformational change. This hypothesis is supported by the observation that the TG-catalyzed activation of HIV-1 PR was completely abolished by spermidine (SPD) which acts as a competitive inhibitor of epsilon-(gamma-glutamyl)lysine formation. Indeed, in the presence of 1mM SPD the formation of the isopeptide was decreased of about 80%. The main products of the TG-catalyzed modification of HIV-1 PR in the presence of SPD were N(1)-mono(gamma-glutamyl)SPD and N(8)-mono(gamma-glutamyl)SPD. Negligible amount of N(1),N(8)-bis(gamma-glutamyl)SPD were found. The significance of these results is discussed with respect to the activation of the protease by post-translational modification and design of potential inhibitors., (2010 Elsevier Inc. All rights reserved.)

Histatin 5 is a cationic salivary peptide with strong candidacidal and bactericidal activity at physiological concentration. In this paper we demonstrate by optical spectroscopy and ESI-IT-MS experiments that a synthetic peptide related to the N-terminus of histatin 5 specifically binds copper ions in vitro and that the complex metal-peptide generates reactive oxygen species at physiological concentration of ascorbate, leading to significant auto-oxidation of the peptide within short reaction time. The oxidative activity of this peptide is associated to the presence of a specific metal binding site present at its N-terminus. The motif is constituted by the amino acid sequence NH(2)-Asp-Ser-His, representing a copper and nickel amino terminal binding site, known as "ATCUN motif". The results of the study suggest that the production of reactive oxygen species can be an intrinsic property of histatin 5 connected to its ability to bind metals.

Reticulons (RTNs) are endoplasmic reticulum-associated proteins widely distributed in plants, yeast, and animals. They are characterized by unique N-terminal parts and a common 200 amino acid C-terminal domain containing two long hydrophobic sequences. Despite their implication in many cellular processes, their molecular structure and function are still largely unknown. In this study, the reticulon family member RTN-1C has been expressed and purified in Escherichia coli and its molecular structure has been analysed by fluorescence and CD spectroscopy in different detergents in order to obtain a good solubility and a relative stability. The isotopically enriched protein has been also produced to perform structural studies by NMR spectroscopy. The preliminary results obtained showed that RTN-1C protein possesses helical transmembrane segments when a membrane-like environment is produced by detergents. Moreover, fluorescence experiments indicated the exposure of tryptophan side chains as predicted by structure prediction programs. We also produced the isotopically labelled protein and the procedure adopted allowed us to plan future NMR studies to investigate the biochemical behaviour of reticulon-1C and of its peptides spanning out from the membrane.

The Azotobacter vinelandii rhodanese is a 31kDa sulfurtransferase protein that catalyzes the transfer of sulfur atom from thiosulfate to cyanide in the detoxification process from cyanide and is able to insert sulfur atom in the iron-sulfur cluster. A study of the uniformly 15N isotopic labeling by high resolution NMR, before obtaining the backbone sequential assignment, has been carried out. The sulfur loaded and the sulfur discharged forms of the enzyme show very similar HSQC spectra with a good spectral dispersion. Few resonances show changes in chemical shift between the two forms. Relaxation parameters T(1), T(2) and 1H-15N NOE of all amide nitrogen atoms, as well as isotope exchange kinetics, show that the two forms exhibit the same global correlation time and hydrodynamic properties. In parallel, essential dynamics studies show that formation and discharging of catalytic cysteine persulfide group has no significant impact on the overall conformation of the protein. These results, taken together, give a clearcut answer to the question if the catalytic mechanism of the enzyme involves a change in the conformation and/or in the mutual orientation of the two domains. On the contrary these results clearly indicate that upon the catalytic mechanism the two domains of the protein behave as a unique fold.

To investigate structural relationship between amphibian and mammalian GSTs the complete amino acid sequence of the major form of glutathione transferase present in toad liver (Bufo bufo) was determined. The enzyme subunit is composed of 210 amino acid residues corresponding to a molecular mass of 24,178 Da. In comparison with the primary structure of amphibian bbGSTP1-1, toad liver GST showed 54% sequence identity. On the other hand, toad liver GST showed about 45-55% sequence identity when compared with other pi class GST and less then 25% identity with GST of other classes. Amino acid residues involved in the H site and in the key and lock structure of the toad enzyme are significantly different from those of bbGSTP1-1 and other mammalian pi class GST. On the basis of its structural and immunological properties the toad liver GST, indicated as bbGSTP2-2, could represent the prototype of a subset of the pi family., (Copyright 2002 Elsevier Science Ltd.)