PSL Chemical Biology Symposia: Recent Progress in Ferroptosis.

This symposium is the 5th PSL (Paris Sciences & Lettres) Chemical Biology meeting (2015, 2016, 2019, 2023, 2024) held at Institut Curie. This initiative originally started at Institut de Chimie des Substances Naturelles (ICSN) in Gif-sur-Yvette, with a strong focus on chemistry. It was then continued at the Institut Curie (2015) covering a larger scope, before becoming the official PSL Chemical Biology meeting. This latest edition hosted around 150 participants and was focused on the burgeoning field of ferroptosis, its mechanism and implications in health and disease. While not initially planned, it was felt that the next large Ferroptosis venue (CSHA, China) would not happen before late 2024. A discussion involving Conrad, Birsoy, Ubellacker, Brabletz and Rodriguez next to lake Como in Italy sponsored by the DKFZ, prompted us to fill in this gap and to organize a Ferroptosis meeting in Paris beforehand.

From desertification to restoration in the Brazilian semiarid region: Unveiling the potential of land restoration on soil microbial properties.

Land desertification poses a significant challenge in the Brazilian semiarid region, encompassing a substantial portion of the country. Within this region, the detrimental effects of human activities, particularly unsuitable anthropic actions, have resulted in diminished vegetation cover and an accelerated rate of soil erosion. Notably, practices such as overgrazing and the conversion of native forests into pasturelands have played a pivotal role in exacerbating the process of land desertification. Ultimately, land desertification results in significant losses of soil organic matter and microbial diversity. To address this pressing issue and contribute to the existing literature, various land restoration practices, such as grazing exclusion, cover crops, and terracing, have been implemented in the Brazilian semiarid. These practices have shown promising results in terms of enhancing soil fertility and restoring microbial properties. Nonetheless, their effectiveness in improving soil microbial properties in the Brazilian semiarid region remains a subject of ongoing study. Recent advances in molecular techniques have improved our understanding of microbial communities in lands undergoing desertification and restoration. In this review, we focus on assessing the effectiveness of these restoration practices in revitalizing soil microbial properties, with a particular emphasis on the soil microbiome and its functions. Through a critical assessment of the impact of these practices on soil microbial properties, our research aims to provide valuable insights that can help mitigate the adverse effects of desertification and promote sustainable development in this ecologically sensitive region.

Contamination risk by heavy metals and enzymatic stoichiometry in agricultural soils under intense use of pesticides.

Soil contamination by heavy metals (HM) from pesticides poses a serious environmental threat, affecting sustainability and agricultural productivity. Soil enzymes are essential for biochemical reactions such as organic matter decomposition and nutrient cycling and are vital for maintaining soil health. However, the effects of HM on soil enzyme activity are not yet well understood. This study examined the impact of HM contamination on enzymatic stoichiometry in regions with intensive pesticide use. We selected flower cultivation areas with 5 years (CA1) and 10 years (CA2) of pesticide exposure and a native forest area (NFA) as a reference during the dry and rainy seasons. We measured Cd, Cu, Mn, Pb, and Zn levels and employed ecological risk indices to assess contamination levels. We also analyzed enzyme activities (arylsulfatase, ß-glucosidase, acid phosphatase, urease) and enzymatic stoichiometry. CA2 exhibited the highest concentrations of Cd, Cu, and Mn in both periods, while Zn was highest in both CA1 and CA2. CA2 had higher values for all indices, indicating significant contamination. Compared with NFA, arylsulfatase activity was lower in cultivated areas during both periods, suggesting decreased soil quality. We found negative correlations between Cu, Mn, Zn, and arylsulfatase, as well as a reduction in urease with Cd; these elements also increased microbial C limitation. Our findings show that continuous pesticide input increases HM levels and that enzyme activity and stoichiometry are effective bioindicator of soil contamination. This study underscores the urgent need for guidelines to protect soils from prolonged HM buildup.

Environmental DNA Sequencing to Monitor Restoration Practices on Soil Bacterial and Archaeal Communities in Soils Under Desertification in the Brazilian Semiarid.

Soils from Brazilian semiarid regions are highly vulnerable to desertification due to their geology, climate, human actions, and intensive land use that contribute to desertification. Therefore, areas under desertification have increased in the Brazilian semiarid region and it has negatively changed the soil bacterial and archaeal communities and their functionality. On the other hand, although restoration strategies are expensive and there are few soils restoration programs, some practices have been applied to restore these soils under desertification. For instance, conservationist practices and grazing exclusion have been strategically implemented, and they created a new altered soil condition for soil microbial communities, boosting soil microbial diversity. Here, we discuss the potential of these restoration strategies to recover the richness and diversity of soil bacterial and archaeal communities that were described through environmental DNA (eDNA) sequencing of soil samples. eDNA sequencing results show that areas where restoration strategies have been applied in regions under desertification in the Brazilian semiarid have increased species richness, diversity, and structure of the bacterial and archaeal community. In addition, network connectivity and functionality of the soil microorganisms have been improved over time. Altogether, we show that management strategies for soil restoration have positive effects on soil microbial communities and these effects can be monitored using the eDNA sequencing approach.

Domestication of Lima Bean (Phaseolus lunatus) Changes the Microbial Communities in the Rhizosphere.

Plants modulate the soil microbiota and select a specific microbial community in the rhizosphere. However, plant domestication reduces genetic diversity, changes plant physiology, and could have an impact on the associated microbiome assembly. Here, we used 16S rRNA gene sequencing to assess the microbial community in the bulk soil and rhizosphere of wild, semi-domesticated, and domesticated genotypes of lima bean (Phaseolus lunatus), to investigate the effect of plant domestication on microbial community assembly. In general, rhizosphere communities were more diverse than bulk soil, but no differences were found among genotypes. Our results showed that the microbial community's structure was different from wild and semi-domesticated as compared to domesticated genotypes. The community similarity decreased 57.67% from wild to domesticated genotypes. In general, the most abundant phyla were Actinobacteria (21.9%), Proteobacteria (20.7%), Acidobacteria (14%), and Firmicutes (9.7%). Comparing the different genotypes, the analysis showed that Firmicutes (Bacillus) was abundant in the rhizosphere of the wild genotypes, while Acidobacteria dominated semi-domesticated plants, and Proteobacteria (including rhizobia) was enriched in domesticated P. lunatus rhizosphere. The domestication process also affected the microbial community network, in which the complexity of connections decreased from wild to domesticated genotypes in the rhizosphere. Together, our work showed that the domestication of P. lunatus shaped rhizosphere microbial communities from taxonomic to a functional level, changing the abundance of specific microbial groups and decreasing the complexity of interactions among them.

Biochar as a carbonaceous material to enhance soil quality in drylands ecosystems: A review.

Drylands are fragile environments that should be carefully managed to improve their quality and functions to achieve sustainable development. Their major problems involve low availability of nutrients and soil organic carbon content. Biochar effect on soil is a joint response of micro to nano sized biochar and soil characteristics. In this review, we attempt to carry out a critical analysis of biochar application to enhance dryland soil quality. Correlating the effects identified from its soil application, we explored the subjects that remains open in the literature. The relation of composition-structure-properties of biochar vary among pyrolysis parameters and biomass sources. Limitations in soil physical quality in drylands, such as low water-holding capacity, can be alleviated by applying biochar at a rate of 10 Mg ha-1 also resulting in beneficial effects on soil aggregation, improved soil porosity, and reduced bulk density. Biochar addition can contribute to the rehabilitation of saline soils, by releasing cations able to displaces sodium in the exchange complex. However, the recovery process of salt-affected soils might be accelerated by the association of biochar with another soil conditioners. This is a promising strategy especially considering the biochar alkalinity and variability in nutrients bioavailability to improve soil fertilization. Further, while higher biochar application rate (>20 Mg ha-1) might change soil C dynamics, a combination of biochar and nitrogen fertilizer can increase microbial biomass carbon in dryland systems. Other aspect of biochar soil application is the economic viability of scale-up production, which is mainly associate to pyrolysis process being biochar production the costliest stage. Nevertheless, the supplying of feedstock might also represent a great input on biochar final costs. Therefore, biochar-based technology is a big opportunity to improve fragile environments such as drylands, integrating sustainable technologies with regional development. Considering the specificity of application area, it might be a model of sustainable agricultural practices protecting the environment in a bioeconomic perspective.

Response of the plant core microbiome to Fusarium oxysporum infection and identification of the pathobiome.

Plant core microbiomes consist of persistent key members that provide critical host functions, but their assemblages can be interrupted by biotic and abiotic stresses. The pathobiome is comprised of dynamic microbial interactions in response to disease status of the host. Hence, identifying variation in the core microbiome and pathobiome can significantly advance our understanding of microbial-microbial interactions and consequences for disease progression and host functions. In this study, we combined glasshouse and field studies to analyse the soil and plant rhizosphere microbiome of cotton plants (Gossypium hirsutum) in the presence of a cotton-specific fungal pathogen, Fusarium oxysporum f. sp. vasinfectum (FOV). We found that FOV directly and consistently altered the rhizosphere microbiome, but the biocontrol agents enabled microbial assemblages to resist pathogenic stress. Using co-occurrence network analysis of the core microbiome, we identified the pathobiome comprised of the pathogen and key associate phylotypes in the cotton microbiome. Isolation and application of some negatively correlated pathobiome members provided protection against plant infection. Importantly, our field survey from multiple cotton fields validated the pattern and responses of core microbiomes under FOV infection. This study advances key understanding of core microbiome responses and existence of plant pathobiomes, which provides a novel framework to better manage plant diseases in agriculture and natural settings.

Tolerance and reduction of chromium by bacterial strains.

Bacteria have potential to tolerate and reduce metals. This study evaluated the potential of selected bacterial strains in tolerating and reducing chromium (Cr). Six bacterial strains (Rhizobium miluonense LCC01, LCC04, LCC05, and LCC69; Rhizobium pusense LCC43; and Agrobacterium deltaense LCC50) showed tolerance to Cr(VI) (16 and 32 µg mL-1), reduction potential of Cr(VI) (from 50 to 80%), and efficiency in producing exopolysaccharides. Rhizobium pusense LCC43 exhibited the highest tolerance (128 µg mL-1), reduction potential of Cr(VI) (from 80 to 100%), and efficiency in producing exopolysaccharides. These results suggested that this strain may have the potential to be used in the bioremediation of soils contaminated with Cr(VI).

Plant growth-promoting mechanisms and genetic diversity of bacteria strains isolated from Brachiaria humidicola and Brachiaria decumbens.

Plant growth-promoting bacteria (PGPB) have received great interest in recent decades. However, PGPB mechanisms remain poorly understood in forage species. We aimed to evaluate roots endophytic and rhizospheric bacteria strains from Brachiaria humidicola and Brachiaria decumbens. The strains were evaluated for biological nitrogen-fixing in saline stress (0 to 10.0 g L-1 of NaCl), N-acyl homoserine lactones and indole-like compounds (ILC) production, the activity of hydrolytic enzymes, and inorganic phosphate solubilization (IPS) under different C sources. The diversity of strains was assessed by BOX-PCR. About 58% of strains were positive for BNF. High salinity levels reduced the growth and BNF. About 58% produced N-acyl homoserine lactones. The ILC was present in 39% of strains. Cellulase, polygalacturonase, pectate lyase, and amylase production were observed in 77, 14, 22, and 25% of strains, respectively. The IPS was observed in 44, 81, and 87% of isolates when glucose, mannitol and sucrose were used, respectively. Comparing two plant species and niches, the strains associated with B. humidicola and root endophytic presented more PGPB mechanisms than others. We found high strain diversity, of which 64% showed similarity lower than 70%. These results can be supporting the bioproducts development to increase forage grasses production in tropical soils.

Probiotic Greek yogurt: effect of the addition of prebiotic fat substitutes on the physicochemical characteristics, probiotic survival, and sensory acceptance.

In this research communication we evaluate the impact of the addition of prebiotic components (inulin, polydextrose, and modified starch, 40 g/l) as fat substitutes on the physicochemical characteristics, probiotic survival, and sensory acceptance of probiotic (Lacticaseibacillus casei 01, 108 CFU/ml) Greek yogurts during storage (7 °C, 28 d). All formulations had probiotic counts higher than 107 CFU/ml during storage and simulated gastrointestinal conditions (SGIC). The prebiotic components increased the probiotic survival to the enteric phase of the SGIC, with inulin producing the most pronounced effect. Inulin addition resulted in products with lower pH values and consistency and higher titratable acidity during storage, with negative impact on the sensory acceptance (flavor, texture, and overall impression) at the end of the storage period. Modified starch addition impacted negatively on the acceptance of the products (appearance, flavor, texture, and overall impression). Polydextrose addition resulted in products with lower consistency, but similar sensory acceptance to the full-fat yogurt. It can be concluded that it is possible to prepare potentially synbiotic Greek yogurts by desorption technique using L. casei as probiotic culture and inulin, polydextrose or modified starch as prebiotic components, with the utilization of polydextrose being advisable.

How deep can ectomycorrhizas go? A case study on Pisolithus down to 4 meters in a Brazilian eucalypt plantation.

Despite the strong ecological importance of ectomycorrhizal (ECM) fungi, their vertical distribution remains poorly understood. To our knowledge, ECM structures associated with trees have never been reported in depths below 2 meters. In this study, fine roots and ECM root tips were sampled down to 4-m depth during the digging of two independent pits differing by their water availability. A meta-barcoding approach based on Illumina sequencing of internal transcribed spacers (ITS1 and ITS2) was carried out on DNA extracted from root samples (fine roots and ECM root tips separately). ECM fungi dominated the root-associated fungal community, with more than 90% of sequences assigned to the genus Pisolithus. The morphological and barcoding results demonstrated, for the first time, the presence of ECM symbiosis down to 4-m. The molecular diversity of Pisolithus spp. was strongly dependent on depth, with soil pH and soil water content as primary drivers of the Pisolithus spp. structure. Altogether, our results highlight the importance to consider the ECM symbiosis in deep soil layers to improve our understanding of fine roots functioning in tropical soils.

Draft genome sequence of cellulose-degrading Bacillus stercoris BHUJPV-SS7 isolated from soil mixed with wood powder.

We report a complete genome of Bacillus stercoris BHUJPV-SS7 isolated from soil which contains 4,299 predicted genes and 4,012 predicted protein-coding genes within its chromosome (4,115,399 bp), and has 43.51% G + C content and a predicted beta-1,4-glucanase (EC 3.2.1.4) gene.

Microdose lithium improves behavioral deficits and modulates molecular mechanisms of memory formation in female SAMP-8, a mouse model of accelerated aging.

Alzheimer's disease (AD) is the most common neuronal disorder that leads to the development of dementia. Until nowadays, some therapies may alleviate the symptoms, but there is no pharmacological treatment. Microdosing lithium has been used to modify the pathological characteristics of the disease, with effects in both experimental and clinical conditions. The present work aimed to analyze the effects of this treatment on spatial memory, anxiety, and molecular mechanisms related to long-term memory formation during the aging process of a mouse model of accelerated aging (SAMP-8). Female SAMP-8 showed learning and memory impairments together with disruption of memory mechanisms, neuronal loss, and increased density of senile plaques compared to their natural control strain, the senescence-accelerated mouse resistant (SAMR-1). Chronic treatment with lithium promoted memory maintenance, reduction in anxiety, and maintenance of proteins related to memory formation and neuronal density. The density of senile plaques was also reduced. An increase in the density of gamma-aminobutyric acid A (GABAA) and α7 nicotinic cholinergic receptors was also observed and related to neuroprotection and anxiety reduction. In addition, this microdose of lithium inhibited the activation of glycogen synthase kinase-3beta (GSK-3ß), the classical mechanism of lithium cell effects, which could contribute to the preservation of the memory mechanism and reduction in senile plaque formation. This work shows that lithium effects in neuroprotection along the aging process are not related to a unique cellular mechanism but produce multiple effects that slowly protect the brain along the aging process.

An Edible Antibacterial Coating Integrating Lytic Bacteriophage Particles for the Potential Biocontrol of Salmonella enterica in Ripened Cheese.

The goal of this research was to create an antibacterial biopolymeric coating integrating lytic bacteriophages against Salmonella enterica for use in ripened cheese. Salmonella enterica is the main pathogen that contaminates food products and the food industry. The food sector still uses costly and non-selective decontamination and disease control methods. Therefore, it is necessary to look for novel pathogen biocontrol technologies. Bacteriophage-based biocontrol seems like a viable option in this situation. The results obtained show promise for food applications since the edible packaging developed (EdiPhage) was successful in maintaining lytic phage viability while preventing the contamination of foodstuff with the aforementioned bacterial pathogen.

Applicability of terahertz spectroscopy in dentistry: a scoping review.

OBJECTIVE: This study aimed to analyze the scope, nature, and extent of the applicability of terahertz (THz) spectroscopy in dentistry. STUDY DESIGN: A scoping review was conducted following the 5-step methodology of Arksey and O'Malley, the PRISMA-ScR checklist, and the Evidence Synthesis Manual of the Joanna Briggs Institute. Electronic literature searches were performed in the PubMed/MEDLINE, Scopus, and Cochrane Library databases, including full-text articles with no specific publication period. The following research question was formulated: "What are the applications of THz spectroscopy in the field of dentistry?" RESULTS: Seventeen laboratory studies were identified, detailing oral and dental applications of THz. In restorative dentistry, 8 investigations sought to identify the properties of human and animal dental tissues and differentiate between healthy and decayed tissue. In oral pathology, 5 articles analyzed the identification of cancer cells in comparison to the identification of these cells in histological or cytological analysis. In biomaterials, 4 papers studied the changes in properties of restorative materials and effects on polymerization. CONCLUSION: While the potential for early diagnosis using THz spectroscopy in dentistry is evident, our findings underscore its limitations. The studies were exclusively conducted in vitro, emphasizing the need for innovative clinical research using intraoral devices. Bridging this gap is essential to unlock the full potential of this noninvasive technology for early diagnosis and informed clinical decision-making.

Unearthing the power of microbes as plant microbiome for sustainable agriculture.

In recent years, research into the complex interactions and crosstalk between plants and their associated microbiota, collectively known as the plant microbiome has revealed the pivotal role of microbial communities for promoting plant growth and health. Plants have evolved intricate relationships with a diverse array of microorganisms inhabiting their roots, leaves, and other plant tissues. This microbiota mainly includes bacteria, archaea, fungi, protozoans, and viruses, forming a dynamic and interconnected network within and around the plant. Through mutualistic or cooperative interactions, these microbes contribute to various aspects of plant health and development. The direct mechanisms of the plant microbiome include the enhancement of plant growth and development through nutrient acquisition. Microbes have the ability to solubilize essential minerals, fix atmospheric nitrogen, and convert organic matter into accessible forms, thereby augmenting the nutrient pool available to the plant. Additionally, the microbiome helps plants to withstand biotic and abiotic stresses, such as pathogen attacks and adverse environmental conditions, by priming the plant's immune responses, antagonizing phytopathogens, and improving stress tolerance. Furthermore, the plant microbiome plays a vital role in phytohormone regulation, facilitating hormonal balance within the plant. This regulation influences various growth processes, including root development, flowering, and fruiting. Microbial communities can also produce secondary metabolites, which directly or indirectly promote plant growth, development, and health. Understanding the functional potential of the plant microbiome has led to innovative agricultural practices, such as microbiome-based biofertilizers and biopesticides, which harness the power of beneficial microorganisms to enhance crop yields while reducing the dependency on chemical inputs. In the present review, we discuss and highlight research gaps regarding the plant microbiome and how the plant microbiome can be used as a source of single and synthetic bioinoculants for plant growth and health.

Short-term restoration practices change the bacterial community in degraded soil from the Brazilian semiarid.

Land degradation by deforestation adversely impacts soil properties, and long-term restoration practices have been reported to potentially reverse these effects, particularly on soil microorganisms. However, there is limited knowledge regarding the short-term effects of restoration on the soil bacterial community in semiarid areas. This study evaluates the bacterial community in soils experiencing degradation (due to slash-and-burn deforestation) and restoration (utilizing stone cordons and revegetation), in comparison to a native soil in the Brazilian semiarid region. Three areas were selected: (a) under degradation; (b) undergoing short-term restoration; and (c) a native area, and the bacterial community was assessed using 16S rRNA sequencing on soil samples collected during both dry and rainy seasons. The dry and rainy seasons exhibited distinct bacterial patterns, and native sites differed from degraded and restoration sites. Chloroflexi and Proteobacteria phyla exhibited higher prevalence in degraded and restoration sites, respectively, while Acidobacteria and Actinobacteria were more abundant in sites undergoing restoration compared to degraded sites. Microbial connections varied across sites and seasons, with an increase in nodes observed in the native site during the dry season, more edges and positive connections in the restoration site, and a higher occurrence of negative connections in the degradation site during the rainy season. Niche occupancy analysis revealed that degradation favored specialists over generalists, whereas restoration exhibited a higher prevalence of generalists compared to native sites. Specifically, degraded sites showed a higher abundance of specialists in contrast to restoration sites. This study reveals that land degradation impacts the soil bacterial community, leading to differences between native and degraded sites. Restoring the soil over a short period alters the status of the bacterial community in degraded soil, fostering an increase in generalist microbes that contribute to enhanced soil stability.

Soil fertility impact on recruitment and diversity of the soil microbiome in sub-humid tropical pastures in Northeastern Brazil.

Soil fertility is key point to pastures systems and drives the microbial communities and their functionality. Therefore, an understanding of the interaction between soil fertility and microbial communities can increase our ability to manage pasturelands and maintain their soil functioning and productivity. This study probed the influence of soil fertility on microbial communities in tropical pastures in Brazil. Soil samples, gathered from the top 20 cm of twelve distinct areas with diverse fertility levels, were analyzed via 16S rRNA sequencing. The soils were subsequently classified into two categories, namely high fertility (HF) and low fertility (LF), using the K-Means clustering. The random forest analysis revealed that high fertility (HF) soils had more bacterial diversity, predominantly Proteobacteria, Nitrospira, Chloroflexi, and Bacteroidetes, while Acidobacteria increased in low fertility (LF) soils. High fertility (HF) soils exhibited more complex network interactions and an enrichment of nitrogen-cycling bacterial groups. Additionally, functional annotation based on 16S rRNA varied between clusters. Microbial groups in HF soil demonstrated enhanced functions such as nitrate reduction, aerobic ammonia oxidation, and aromatic compound degradation. In contrast, in the LF soil, the predominant processes were ureolysis, cellulolysis, methanol oxidation, and methanotrophy. Our findings expand our knowledge about how soil fertility drives bacterial communities in pastures.

Grazing exclusion-induced changes in soil fungal communities in a highly desertified Brazilian dryland.

Soil desertification poses a critical ecological challenge in arid and semiarid climates worldwide, leading to decreased soil productivity due to the disruption of essential microbial community processes. Fungi, as one of the most important soil microbial communities, play a crucial role in enhancing nutrient and water uptake by plants through mycorrhizal associations. However, the impact of overgrazing-induced desertification on fungal community structure, particularly in the Caatinga biome of semiarid regions, remains unclear. In this study, we assessed the changes in both the total fungal community and the arbuscular mycorrhizal fungal community (AMF) across 1. Natural vegetation (native), 2. Grazing exclusion (20 years) (restored), and 3. affected by overgrazing-induced degradation (degraded) scenarios. Our assessment, conducted during both the dry and rainy seasons in Irauçuba, Ceará, utilized Internal Transcribed Spacer (ITS) gene sequencing via Illumina® platform. Our findings highlighted the significant roles of the AMF families Glomeraceae (∼71% of the total sequences) and Acaulosporaceae (∼14% of the total sequences) as potential key taxa in mitigating climate change within dryland areas. Moreover, we identified the orders Pleosporales (∼35% of the total sequences) and Capnodiales (∼21% of the total sequences) as the most abundant soil fungal communities in the Caatinga biome. The structure of the total fungal community differed when comparing native and restored areas to degraded areas. Total fungal communities from native and restored areas clustered together, suggesting that grazing exclusion has the potential to improve soil properties and recover fungal community structure amid global climate change challenges.

Structural characterization of the human DjC20/HscB cochaperone in solution.

J-domain proteins (JDPs) form a very large molecular chaperone family involved in proteostasis processes, such as protein folding, trafficking through membranes and degradation/disaggregation. JDPs are Hsp70 co-chaperones capable of stimulating ATPase activity as well as selecting and presenting client proteins to Hsp70. In mitochondria, human DjC20/HscB (a type III JDP that possesses only the conserved J-domain in some region of the protein) is involved in [FeS] protein biogenesis and assists human mitochondrial Hsp70 (HSPA9). Human DjC20 possesses a zinc-finger domain in its N-terminus, which closely contacts the J-domain and appears to be essential for its function. Here, we investigated the hDjC20 structure in solution as well as the importance of Zn+2 for its stability. The recombinant hDjC20 was pure, folded and capable of stimulating HSPA9 ATPase activity. It behaved as a slightly elongated monomer, as attested by small-angle X-ray scattering and SEC-MALS. The presence of Zn2+ in the hDjC20 samples was verified, a stoichiometry of 1:1 was observed, and its removal by high concentrations of EDTA and DTPA was unfeasible. However, thermal and chemical denaturation in the presence of EDTA led to a reduction in protein stability, suggesting a synergistic action between the chelating agent and denaturators that facilitate protein unfolding depending on metal removal. These data suggest that the affinity of Zn+2 for the protein is very high, evidencing its importance for the hDjC20 structure.