BioTech最新文献

The synthesis of enantiomeric forms of D-amino acids can be achieved by a two-step "hydantoinase process" based on the sequential catalysis of substrates by specific enzymes, D-carbamoylase and D-hydantoinase. Here, we describe the structural features of D-carbamoylase from Pseudomonas, the encoded gene of which was chemically synthesized and cloned into Escherichia coli. A significant fraction of the overexpressed recombinant protein forms insoluble inclusion bodies, which are partially converted to a soluble state upon treatment with N-lauroylsarcosine or upon incubation of cells at 28 °C. Purified His-tagged protein exhibits the highest activity towards N-carbamoyl-D-alanine and N-carbamoyl-D-tryptophan. Comprehensive virtual analysis of the interactions of bulky carbamylated amino acids with D-carbamoylase provided valuable information. Molecular docking analysis revealed the location of the substrate binding site in the three-dimensional structure of D-carbamoylase. Molecular dynamics simulations showed that the binding pocket of the enzyme in complex with N-carbamoyl-D-tryptophan was stabilized within 100 nanoseconds. The free energy data showed that Arg176 and Asn173 formed hydrogen bonds between the enzyme and substrates. The studies of D-carbamoylases and the properties of our previously obtained D-hydantoinase suggest the possibility of developing a harmonized biotechnological process for the production of new drugs and peptide hormones.

Cadherin-catenin cell-cell adhesion complexes, composed of cadherin, β-catenin or plakoglobin, and α-catenin (α-cat) molecules, are crucial for maintaining cell-cell contact and are commonly referred to as "adherens junctions (AJs)." Inactivating this system leads to loss of cell-cell contact and developmental arrest in early embryos. However, it remains unclear whether the loss of cell-cell contact affects the differentiation of embryonic cells. In this study, we explored the use of a murine embryonal carcinoma cell line, P19, as an in vitro model for early embryogenesis. P19 cells easily form embryoid bodies (EBs) and are susceptible to cellular differentiation in response to retinoic acid (RA) and teratoma formation. Using CRISPR/Cas9 technology to disrupt the endogenous α-cat gene in P19 cells, we generated α-cat knockout (KO) cells that exhibited a loss of cell-cell contact. When cultivated on non-coated dishes, these α-cat KO cells formed EBs, but their structures were labile. In the RA-containing medium, the α-cat KO EBs failed to produce differentiated cells on their outer layer and continued to express SSEA-1, an antigen specific to pluripotent cells. Teratoma formation assays revealed an absence of overt differentiated cells in tumors derived from α-cat KO P19 cells. Aggregation assays revealed the inability of the KO cells to colonize into the zona pellucida-denuded 8-cell embryos. These findings suggest that the AJs are essential for promoting the early stages of cellular differentiation and for the colonization of early-developing embryos.

In Tunisia, the date industry generates a large quantity of waste, raising environmental concerns. However, dates are rich in sugars, which offer a renewable source of nutrients for various applications. In this study, sugar extraction from two low-grade pitted date fruits (Alig and Kentichi) under ultrasound, was optimized using full factorial design. At 40 °C, for20 min, and with a liquid-to-solid ratio of 10 mL/g, the optimum sugar contents were 60.87% and 50.79% for the varieties Alig and Kentichi, respectively. The date extracts were chemically analyzed, revealing low fat and protein contents, but significant polyphenol and mineral contents in both varieties. HPLC-IR analysis revealed more inverted sugars (glucose and fructose) in the Alig variety and more sucrose in the Kentichi variety. FTIR and SEM analysis showed the efficiency of the ultrasonic treatment of the biomass in terms of improving mass transfer diffusion through ultrasonic cavitation. Thus, ultrasound-assisted extraction constitutes an effective method for the recovery of sugar from date waste.

Currently, starter cultures for fermenting plant-based beverages are not widely available commercially, but producers can use starter cultures for dairy products. Therefore, the aim of this study was to determine the physicochemical, rheological, antioxidant and sensory properties of oat beverages with/without pectin fermented by four different dairy starter cultures. The use of a mono-starter with Lactobacillus bulgaricus or Sreptococcus thermophilus allows for the efficient use of glucose, and more lactic acid is accumulated. The beverage with L. bulgaricus is characterised by high adhesion, syneresis and low cohesiveness, and it has high antioxidant activity and a low sensory profile. Using starter with L. bulgaricus, S. thermophilus and some Lactococcus for fermentation yields a product with high sensory capacity, forming a high-viscosity beverage matrix with low syneresis, high water retention, chewy texture and stickiness. It has been observed that the absence of lactococci and the presence of Lactobacillus casei, L. Rhamnosus and L. paracasei in the starter yields a product with high antioxidant activity, especially in the presence of pectin. The use of pectin significantly improves the viscosity and textural properties of oat yoghurt, enhancing the drink's flavour and giving it body. For many reasons, the use of different commercial starters in the dairy industry results in different viscosities of oat fermented beverages, forming a matrix with different textural, sensory and antioxidant properties.

Viruses and viroids pose a significant challenge in citriculture, and their control is crucial for plant health. This study evaluated the effectiveness of in vitro thermotherapy combined with a meristem tip culture for eliminating citrus exocortis viroid (CEVd) and hop stunt viroid (HSVd) from a new limonime hybrid (Citrus x limon var. limon x Citrus latifolia var. latifolia). The elimination success was confirmed by RT-PCR assays. The in vitro elimination rate for CEVd during the shoot proliferation stage (43%) was higher than for HSVd (21%). Accordingly, in the subsequent rooting stage, the in vitro elimination rate for CEVd (50%) was higher than for HSVd (33%). Successful CEVd and HSVd eradication at a 100% rate was confirmed in the ex vitro acclimatized plants in the greenhouse. The study also established an efficient micropropagation protocol. The optimal treatment for in vitro shoot induction was 0.5-2 mg L^-1 benzyladenine (BA) + 0.5 mg L^-1 gibberellic acid (GA₃) + 0.25 mg L^-1 naphthalene acetic acid (NAA), while for shoot elongation, it was 0.5 mg L^-1 BA + 0.5 mg L^-1 kinetin (KIN) + 0.5 mg L^-1 GA₃ + 0.25 mg L^-1 NAA. Rooting was best promoted by 1 mg L^-1 NAA. This study provides valuable insights for the mass production of viroid-free propagation material in this new lemon x lime hybrid, contributing to the conservation of genetic resources in citrus breeding programs through the combined application of in vitro thermotherapy and an in vitro meristem tip culture, a novel and highlighted achievement reported for the first time in this study.

Chronic wounds result from the body's inability to heal, causing pain, pathogen entry, limited treatment options, and societal burden. Diabetic foot ulcers are particularly challenging, often leading to severe complications like leg amputation. A clinical study tested AMNIODERM+^®, a new device with a lyophilized human amniotic membrane (HAM), on chronic diabetic foot ulcers. Participants had diabetic neuropathic or neuroischemic leg wounds (2-16 cm²) unhealed by 20% after six weeks of standard care. This study showed significant wound healing improvements with AMNIODERM+^®. The median wound size reduction after 12 weeks was 95.5%, far exceeding the null hypothesis of 20% change. Additionally, 65% of patients achieved complete ulceration healing, surpassing the 50% efficacy requirement. The median time to full closure was 11.4 weeks, with the proportion of completely healed patients rising progressively, reaching 55% by week 11. These findings, from the clinical trial "Freeze-dried amniotic membrane in the treatment of nonhealing wounds", suggest AMNIODERM+^® as a promising future treatment for chronic diabetic foot ulcers. The published results were obtained as part of a clinical trial entitled "Freeze-dried amniotic membrane in the treatment of nonhealing wounds: a single-arm, retrospectively-perspective clinical trial", EUDAMED Nr. CIV-SK-22-10-041146.

Power scarcity and pollution can be overcome with the use of green energy forms like ethanol, biogas, electricity, hydrogen, etc., especially energy produced from renewable and industrial feedstocks. In hilly areas, pine needles are the most abundant biomass that has a low possibility of valorization due to high lignin content. On the other hand, anaerobic digestion (AD) of lignin and animal waste has low biogas yield due to poor conductivity. This study focuses on the simultaneous production of biogas and electricity through the co-digestion of cow dung and pine needles. The digester was initially established and stabilized in the lab to ensure a continuous supply of inoculum throughout the experiment. The optimization process involved the determination of an ideal cow dung-to-water ratio and selecting the appropriate conductive material that can enhance the energy generation from the feedstock. Afterward, both batch and continuous anaerobic digestion experiments were conducted. The results revealed that the addition of powdered graphite (5 mM), activated charcoal (15 mM), and biochar (25 mM) exhibited maximum voltage of 0.71 ± 0.013 V, 0.56 ± 0.013 V, and 0.49 ± 0.011 V on the 30th, 25th and 20th day of AD, respectively. The batch experiment showed that 5 mM graphite powder enhanced electron transfer in the AD process and generated a voltage of 0.77 ± 0.014 V on the 30th day, indicating an increase of ~1.5-fold as compared to the control (0.56 ± 0.019 V). The results from the continuous AD process showed that the digester with cow dung, pine needle, and a conductive material in combination exhibited the maximum voltage of 0.76 ± 0.012 V on the 21st day of AD, while the digester with cow dung only exhibited a maximum voltage of 0.62 ± 0.015 V on the 22nd day of AD, representing a 1.3-fold increase over the control. Furthermore, the current work used discarded plastic items and electrodes from spent batteries to emphasize waste management and aid in attaining sustainable energy and development goals.

Biobanking plays a pivotal role in biomedical research by providing standardized processing, precise storing, and management of biological sample collections along with the associated data. Effective data management is a prerequisite to ensure the integrity, quality, and accessibility of these resources. This review provides a current landscape of data management in biobanking, discussing key challenges, existing strategies, and potential future directions. We explore multiple aspects of data management, including data collection, storage, curation, sharing, and ethical considerations. By examining the evolving technologies and methodologies in biobanking, we aim to provide insights into addressing the complexities and maximizing the utility of biobank data for research and clinical applications.

This study introduces the intelligent learning engine (ILE) optimization technology, a novel approach designed to revolutionize screening processes in bioinformatics, cheminformatics, and a range of other scientific fields. By focusing on the efficient and precise identification of candidates with desirable characteristics, the ILE technology marks a significant leap forward in addressing the complexities of candidate selection in drug discovery, protein classification, and beyond. The study's primary objective is to address the challenges associated with optimizing screening processes to efficiently select candidates across various fields, including drug discovery and protein classification. The methodology employed involves a detailed algorithmic process that includes dataset preparation, encoding of protein sequences, sensor nucleation, and optimization, culminating in the empirical evaluation of molecular activity indexing, homology-based modeling, and classification of proteins such as G-protein-coupled receptors. This process showcases the method's success in multiple sequence alignment, protein identification, and classification. Key results demonstrate the ILE's superior accuracy in protein classification and virtual high-throughput screening, with a notable breakthrough in drug development for assessing drug-induced long QT syndrome risks through hERG potassium channel interaction analysis. The technology showcased exceptional results in the formulation and evaluation of novel cancer drug candidates, highlighting its potential for significant advancements in pharmaceutical innovations. The findings underline the ILE optimization technology as a transformative tool in screening processes due to its proven effectiveness and broad applicability across various domains. This breakthrough contributes substantially to the fields of systems optimization and holds promise for diverse applications, enhancing the process of selecting candidate molecules with target properties and advancing drug discovery, protein classification, and modeling.

Seven auxin-producing endophytic bacterial strains (Azospirillum spp., Methylobacterium symbioticum, Bacillus spp.), and two different combinations of these strains were used to verify their influence on tomato during germination and development in hydroponic conditions where, as a novelty for Canestrino di Lucca cultivar, endophytic bacteria were inoculated. To emphasize the presence of bacterial auxins in roots and stems of seedlings, both in situ staining qualitative assessment and quantitative analysis were carried out. Moreover, hypogeal and epigeal growth of the plantlets were measured, and correlation analyses were conducted to examine the relationship between the amount of indolacetic acid (IAA) produced by the bacterial strains and root and stem parameters. Plantlets treated with microbial inoculants showed a significant increase in the survival rate compared to the control treatment. The best results as IAA producers were from Azospirillum baldaniorum Sp245 and A. brasilense Cd, which also induced significant root growth. On the other hand, Bacillus amyloliquefaciens and B. licheniformis induced the best rates in stem growth. These findings highlight the potential for using endophytic bacterial strains in a hydroponic co-cultivation system that enables inoculating plantlets, at an early stage of growth (5 days old).