Synthesizing Amino Acids Modified with Reactive Carbonyls in Silico to Assess Structural Effects Using Molecular Dynamics Simulations.

Protein carbonylation by reactive aldehydes derived from lipid peroxidation leads to cross-linking, oligomerization, and aggregation of proteins, causing intracellular damage, impaired cell functions, and, ultimately, cell death. It has been described in aging and several age-related chronic conditions. However, the basis of structural changes related to the loss of function in protein targets is still not well understood. Hence, a route to the in silico construction of new parameters for amino acids carbonylated with reactive carbonyl species derived from fatty acid oxidation is described. The Michael adducts for Cys, His, and Lys with 4-hydroxy-2-nonenal (HNE), 4-hydroxy-2-hexenal (HHE), and a furan ring form for 4-Oxo-2-nonenal (ONE), were built, while malondialdehyde (MDA) was directly attached to each residue. The protocol describes details for the construction, geometry optimization, assignment of charges, missing bonds, angles, dihedral angles parameters, and its validation for each modified residue structure. As a result, structural effects induced by the carbonylation with these lipid derivatives have been measured by molecular dynamics simulations on different protein systems such as the thioredoxin enzyme, bovine serum albumin and the membrane Zu-5-ankyrin domain employing root-mean-square deviation (RMSD), root mean square fluctuation (RMSF), structural secondary prediction (DSSP) and the solvent-accessible surface area analysis (SASA), among others.

Molecular dynamics of structural effects of reactive carbonyl species derivate of lipid peroxidation on bovine serum albumin.

BACKGROUND: Serum albumin is the most abundant protein in the Mammalia blood plasma at where plays a decisive role in the transport wide variety of hydrophobic ligands. BSA undergoes oxidative modifications like the carbonylation by the reactive carbonyl species (RCSs) 4-hydroxy-2-nonenal (HNE), 4 hydroxy-2-hexenal (HHE), malondialdehyde (MDA) and 4-oxo-2-nonenal (ONE), among others. The structural and functional changes induced by protein carbonylation have been associated with the advancement of neurodegenerative, cardiovascular, metabolic and cancer diseases. METHODS: To elucidate structural effects of protein carbonylation with RCSs on BSA, parameters for six new non-standard amino acids were designated and molecular dynamics simulations of its mono­carbonylated-BSA systems were conducted in the AMBER force field. Trajectories were evaluated by RMSD, RMSF, PCA, RoG and SASA analysis. RESULTS: An increase in the conformational instability for all proteins modified with local changes were observed, without significant changes on the BSA global three-dimensional folding. A more relaxed compaction level and major solvent accessible surface area for modified systems was found. Four regions of high molecular fluctuation were identified in all modified systems, being the subdomains IA and IIIB those with the most remarkable local conformational changes. Regarding essential modes of domain movements, it was evidenced that the most representatives were those related to IA subdomain, while IIIB subdomain presented discrete changes. CONCLUSIONS: RCSs induces local structural changes on mono­carbonylated BSA. Also, this study extends our knowledge on how carbonylation by RCSs induce structural effects on proteins.

Cysteine carbonylation with reactive carbonyl species from lipid peroxidation induce local structural changes on thioredoxin active site.

Protein oxidative modifications with reactive carbonyl species (RCS) is directly linked to metabolic processes in premature aging, cancer, neurodegenerative and infectious diseases. RCS as 4-Hydroxy-2-nonal (HNE), 4-Hydroxy-2-hexenal (HHE), 4-Oxo-2-nonenal (ONE) and Malondialdehyde (MDA) attack nucleophilic amino acids residues forming irreversible adducts with proteins as Thioredoxins (Trx). This is a class of small thiol oxide-reductases playing a key role in redox signaling and oxidative stress responses in mammals. Although proteomic studies have identified to Cys-32 residue as a target of HNE attack that inhibit its enzymatic activity, how this carbonylation affects its structure and dynamic behavior at the atomic level is unknown. Even more, the molecular bases for the atomistic behavior of these modified proteins have not been completely understood. We present molecular dynamics simulations of Trx-modified with four different RCS to analyze its global and local structural effects. For this, parameters supported in the AMBER force fields were built and validated for three non-natural cysteine residues modified with HHE, ONE and MDA. Results obtained showed a slight change in the global conformational stability of Trx modified with HNE and MDA, establishing that all modified proteins presented local regions of high mobility in the modified catalytic site and some regions far from the modification area. In addition, essential domain movement modes evidences that proteins modified with the RCS assayed induce changes in conformational flexibility. Finally, these data showed that the given conformational changes did not caused global changes in proteins but rather localized changes in particular regions.

Protein Susceptibility to Peroxidation by 4-Hydroxynonenal in Hereditary Hemochromatosis.

Iron overload caused by hereditary hemochromatosis (HH) increases free reactive oxygen species that, in turn, induce lipid peroxidation. Its 4-hydroxynonenal (HNE) by-product is a well-established marker of lipid peroxidation since it reacts with accessible proteins with deleterious consequences. Indeed, elevated levels of HNE are often detected in a wide variety of human diseases related to oxidative stress. Here, we evaluated HNE-modified proteins in the membrane of erythrocytes from HH patients and in organs of Hfe-/- male and female mice, a mouse model of HH. For this purpose, we used one- and two-dimensional gel electrophoresis, immunoblotting and MALDI-TOF/TOF analysis. We identified cytoskeletal membrane proteins and membrane receptors of erythrocytes bound to HNE exclusively in HH patients. Furthermore, kidney and brain of Hfe-/- mice contained more HNE-adducted protein than healthy controls. Our results identified main HNE-modified proteins suggesting that HH favours preferred protein targets for oxidation by HNE.

Elemental mercury accumulation in Eichhornia crassipes (Mart.) Solms-Laubach.

The aquatic macrophyte Eichhornia crassipes has great potential for the control of Hg pollution in the environment. The aim of this study was to investigate the capability of E. crassipes to accumulate elemental mercury (Hg0). The plants were exposed for 30 days to 5, 10, 20, 40, and 80 mg of Hg0 in a 1-L Hoagland medium with the Hg0 settled at the bottom of the flask. The roots of the plants did not touch the mercury during the treatment. After exposure, the total Hg (T-Hg) concentrations in the roots, leaves, and stems were measured using a direct mercury (Hg) analyzer. The highest concentrations were found at 80 mg Hg0 treatment in the roots, leaves, and stems, in that order. The translocation factor indicated a poor capability of Hg to translocate from the roots to the shoots. The relative growth and the root-length inhibition measurements showed that the differences between Hg0 treatments were not significant. In addition, the treatments negatively affected the chlorophyll concentration. The carotenoid content was found to be significantly different at 20 and 40 mg of Hg0 in 1 L. Regarding the carbonyl index in root proteins, significant differences compared to control were found at the highest Hg treatment. Based on these results, it was shown that E. crassipes is able to take up elemental Hg from Hoagland medium. However, the Hg0 treatments did not show a strong stress-response activation mechanism in the evaluated plant tissues.

Proteomic Changes in Paspalum fasciculatum Leaves Exposed to Cd Stress.

(1) Background: Cadmium is a toxic heavy metal that is widely distributed in water, soil, and air. It is present in agrochemicals, wastewater, battery waste, and volcanic eruptions. Thus, it can be absorbed by plants and enter the trophic chain. P. fasciculatum is a plant with phytoremediation capacity that can tolerate Cd stress, but changes in its proteome related to this tolerance have not yet been identified. (2) Methods: We conducted a quantitative analysis of the proteins present in P. fasciculatum leaves cultivated under greenhouse conditions in mining soils doped with 0 mg kg-1 (control), 30 mg kg-1, or 50 mg kg-1. This was carried out using the label-free shotgun proteomics technique. In this way, we determined the changes in the proteomes of the leaves of these plants, which allowed us to propose some tolerance mechanisms involved in the response to Cd stress. (3) Results: In total, 329 variable proteins were identified between treatments, which were classified into those associated with carbohydrate and energy metabolism; photosynthesis; structure, transport, and metabolism of proteins; antioxidant stress and defense; RNA and DNA processing; and signal transduction. (4) Conclusions: Based on changes in the differences in the leaf protein profiles between treatments, we hypothesize that some proteins associated with signal transduction (Ras-related protein RABA1e), HSPs (heat shock cognate 70 kDa protein 2), growth (actin-7), and cellular development (actin-1) are part of the tolerance response to Cd stress.

Carbonylation induced by antibiotic and pesticide residues on casein increases its IgE binding and allergenicity.

This work aimed to evaluate the effect of carbonylation induced by tetracyclines, ß-lactams, fluoroquinolones, and pyrethroids in caseins of bovine origin on their immunoreactivity and allergenicity. Using a spectrophotometric method, ELISA, dot-blot, and an IgE-mediated milk allergy mouse model, we confirmed that antibiotics and pesticides at their maximum residue limit, promoted the in vitro carbonylation of caseins (among 5.0 ± 0.01 and 67.5 ± 0.70 nmol of carbonyl/mg of protein); furthermore, carbonylations greater than 19 nmol significantly increase the in vitro IgE immunoreactivity of caseins (average OD among 0.63-1.50) regarding the negative control (average OD: 0.56). On the other hand, sensitized mice exposed to oxidized caseins showed increased clinical scores (2-5), positive skin tests, and footpad swelling (0.28-0.59 mm) compared to the negative control (1-2; negative skin tests; 0.1 mm, respectively), denoting increased allergenicity. These results suggest that casein carbonylation increases their IgE immunoreactivity and allergenicity, a fact that could be explained by the resistance to the digestion promoted by carbonylation and by conformational changes in the random coil casein structure, which can expose cryptic epitopes or neoepitopes.

Quantification of matrix metalloproteinases MMP-8 and MMP-9 in gingival overgrowth.

BACKGROUND: Matrix metalloproteinases (MMPs) are proteolytic enzymes involved in extracellular matrix remodeling of all body tissues, including oral tissues such as gingival tissue. Expression levels of MMPs are widely studied as important biomarkers for explaining the biochemical mechanisms and evolution of many oral diseases. OBJECTIVE: Demonstrate the sensitivity, reproducibility, repeatability, and robustness of the dot blot assay for the relative quantification of MMP-8 and MMP-9 expression levels in patients with GO associated with orthodontic treatment. METHODS: A validated dot blot assay was used to compare the relative expression levels of MMP-8 and MMP-9 in gingival samples. Methodological variability, reproducibility, sensitivity and robustness were determined with the use of control samples from healthy donors (G1). Next, expression levels were measured in gingival tissue from patients with mild and moderate gingival overgrowth associated with orthodontic treatment (G3 and G4) and patients without gingival overgrowth but with a history of using orthodontic appliances (G2). RESULTS: Dot blot assay demonstrated that MMP-8 and MMP-9 expression levels were higher in patients with gingival overgrowth and distinguished those with moderate clinical grade (G4) from those with mild overgrowth (G3). In addition, patients with a history of orthodontic treatment showed similar expression levels to the control group two years after removing orthodontic appliances. CONCLUSIONS: With the assay used, we were able to detect differences in MMP-8 and MMP-9 expression in patients with different levels of severity of gingival overgrowth. Dot blot could be used to measure MMPs during the onset and progression of gingival overgrowth.

Erythrocyte membrane proteins involved in the immune response to Plasmodium falciparum and Plasmodium vivax infection.

Invasion of Plasmodium into the red blood cell involves the interactions of a substantial number of proteins, with red cell membrane proteins as the most involved throughout the process from entry to exit. The objective of this work was to identify proteins of the human erythrocyte membrane capable of generating an antigenic response to P. falciparum and P. vivax infection, with the goal of searching for new molecular targets of interest with an immunological origin to prevent Plasmodium infection. To identify these proteins, an immunoproteomic technique was carried out in four stages: protein separation (electrophoresis), detection of antigenic proteins (western blotting), identification of proteins of interest (mass spectrometry), and interpretation of the data (bioinformatic analysis). Four proteins were identified from extracts of membrane proteins from erythrocytes infected with P. falciparum: Spectrin, Ankyrin-1, Band 3 and band 4.2, and a single protein was identified from erythrocytes infected with P. vivax: Band 3. These results demonstrate that modifications in the red blood cell membrane during infection with P. falciparum and P. vivax can generate an immune response, altering proteins of great structural and functional importance.

Protein carbonylation associated with nickel liberation in orthodontic gingival overgrowth.

OBJECTIVE: To determine nickel levels and their impact on protein carbonylation in gum samples from patients with gingival overgrowth by orthodontic treatment. DESIGN: A retrospective observational study with 33 patients divided into three groups. Group 1 patients with gingival overgrowth by orthodontic appliances; group 2 patients without gingival overgrowth but with a history of orthodontic treatment; group 3 patients without overgrowth and history of orthodontic appliances. Nickel level in gingiva samples was measured by atomic absorption while protein carbonylation was determined by Western Blot. Furthermore, three proteins were identified in carbonylated protein bands by mass spectrometry. RESULTS: Statistically significant differences (p < 0,05) in tissue nickel levels among groups were established (nickel levels group 1: 1.33 ± 1.52; group 2: 0.33 ± 0.44; group 3: 0.20 ± 0.22 µg Ni/g tissue). Protein carbonylation was higher in patients with gingival enlargement (group 1) and history of appliance use (group 2) than controls (group 3). It was observed that band A of the Western blots presented the highest intensity (Rf 0.23) with an average intensity of 4.133.830 ± 1.958.569 for group 1; 4.420.146 ± 1.594.679 for group 2 and 2.110. 727 ± 1.640.721 for group 3. Also, the proteins Teneurin-4, Bromodomain adjacent to zinc finger domain protein 2B, Lysine-specific demethylase 5B, and Serum albumin, were identified from oxidized bands. CONCLUSIONS: The gum of patients with gingival overgrowth by orthodontic appliances contains higher nickel residues and carbonylation of its proteins.

Residues of tetracyclines and ß-lactams antibiotics induce carbonylation of chicken breast.

Background: Worldwide, chicken meat is widely consumed due to its low cost, high nutritional value and non-interference with religious or cultural beliefs. However, during animal husbandry chickens are exposed to many chemical substances, including tetracyclines and ß-lactams, which are used to prevent and cure several infections. Some residues of these compounds may bioaccumulate and be present in chicken meat after slaughtering, promoting oxidative reactions. Methods: In order to evaluate in vitro carbonylation induced by tetracyclines and ß-lactams residues, a proteomic approach was used. For this, chicken muscle was individually contaminated with tetracyclines (tetracycline, chlortetracycline, oxytetracycline, and doxycycline) and ß-lactams (ampicillin, benzathine penicillin, dicloxacillin and oxacillin) at 0.5, 1.0 and 1.5 times their maximum residue level (MRL). Then, sarcoplasmic, myofibrillar and insoluble proteins were extracted and their content were measured using the Bradford method. Protein carbonylation was measured using the 2,4-Dinitrophenylhydrazine alkaline method. Results: Residues of tetracyclines and ß-lactams induced in vitro carbonylation on sarcoplasmic, myofibrillar and insoluble proteins even at 0.5MRL concentrations ( p<0.05). When comparing the carbonylation induced by both antibiotics no differences were found ( p>0.05). Variables such as the partition coefficient (log P) and the concentration of these antibiotics showed a high correlation with the oxidative capacity of tetracyclines and ß-lactams on chicken breast proteins. Conclusions: This study shows that the presence of tetracyclines and ß-lactams residues at MRLs concentrations promotes in vitro carbonylation on chicken breast proteins. Our results provide important insights about the impact of antibiotics on the integrity of meat proteins intended for human consumption.

Fluoroquinolone antibiotics and organophosphate pesticides induce carbonylation on Eisenia fetida muscle proteins.

Protein carbonylation is an irreversible oxidative modification that has been associated with a decrease in the quality and nutritional value of products of animal origin. Generally, the carbonylation is attributed to processes of slaughter, processing, and cold storage of products. However, in vitro studies have shown that fluoroquinolone and organophosphate pesticides residues at their maximum residue limits (MRL) can promote carbonylation of animal proteins. Though, this effect on in vivo conditions has not yet been evaluated. Thus, Eisenia fetida was chosen as a model to assay their oxidant effect. For this, adult earthworms were exposed to artificial soil contaminated with ciprofloxacin, danofloxacin, fenthion, and diazinon at three concentrations (0.5, 1.0, and 1.5 MRL) for 28 days. Then, these were purged and sacrificed to obtain the muscle region between the anus and preclitellum. The muscle samples were cold macerated to obtain muscle proteins, which were used for protein quantification, determination of carbonyl levels, and carbonyl protein profiles employing Bradford, Dot-blot, and Western blot assays, respectively. The results showed that at each concentration assayed, all pollutants induced significant carbonylation respect to control (p < 0.05). Additionally, mass spectrometry-based analysis (MALDI-TOF/TOF) identified actin as the protein most susceptible to carbonylation promoted by these substances. Therefore, these findings show for the first time the oxidant power of fluoroquinolones and organophosphates pesticides at MRLs concentrations on muscle proteins under in vivo conditions. Fact causes concern due to the homology of muscle proteins in eukaryotes, which allow to hypothesize that this effect could also be experienced by proteins from food-producing animals in the same way that observed in in vitro studies.

Peptidomics and proteomics data of oxidised peptides from in vitro gastrointestinal digestion of chicken breast exposed to chlorpyrifos.

Chlorpyrifos (CPF) is an organophosphorus pesticide used in poultry to prevent and/or kill insects and such as preserving agents of poultry feed. Exposure continues to CPF can promote its accumulation at trace concentrations in animal tissue. The toxicological effects of these residues (carcinogenicity, genotoxicity, and neurological disorders) have been related to oxidative stress. Nevertheless, it is still unknown if these trace concentrations might promote oxidative stress in muscle proteins since chicken meat proteins are susceptible to undergo oxidation reactions. Moreover, protein oxidation has been related to a decrease in the nutritional value in of meat. To investigate the oxidative effect of CPF on chicken breast proteins, peptidomics and proteomics analysis were used. For this, chicken breast samples were exposed to CPF and subjected to simulated gastrointestinal digestion. The identification of oxidized peptides from digested and undigested proteins were performed by LC MS/MS (nanoESI qQTOF). Prior to mass analyses undigested proteins were trypsinated. Data were analysed using MASCOT and ProteinPilot v 4.5 software. In this study, 90 and 107 oxidized peptides from digested proteins of control and exposed samples were identified, respectively. These peptides corresponding to 12 oxidized proteins. Meanwhile, 260 and 324 oxidized peptides from undigested proteins (control and exposed samples) were identified, which corresponding to 19 and 17 proteins, respectively. Collagen was protein more susceptible to oxidation promoted by chlorpyrifos in digested and undigested proteins. Identification of these oxidized proteins from simulated digestion provides an important insight about the impact of substances like certain veterinary drugs at trace concentrations on the nutritional value of chicken breast meat.

In vitro oxidation promoted by chlorpyrifos residues on myosin and chicken breast proteins.

Organophosphate pesticides are frequently used to eliminate or prevent insects in poultry. However, their residues may continue in meat after slaughtering. In this study, proteomics and peptidomics approaches were used to evaluate their oxidative impact on myosin and chicken breast proteins under in vitro conditions. Myosin protein was exposed to diazinon and chlorpyrifos showing an increase in its oxidation by increasing times, especially with chlorpyrifos. Then, chicken breast was contaminated with chlorpyrifos to evaluate carbonylation and the effect of simulated gastrointestinal digestion. Proteins were isolated using size-exclusion-chromatography and identified by mass spectrometry in tandem. Myosin, ß-enolase, CK-M-type and actin were identified as main proteins susceptible to oxidation. Also, oxidised peptides obtained before and after simulated gastrointestinal digestion were identified. Collagen peptides the most susceptible to oxidation. These results suggest that the presence of chlorpyrifos residues on meat could have a negative effect on its final quality and nutritional value.

AMBER parameters and topology data of 2-pentylpyrrole adduct of arginine with 4-hydroxy-2-nonenal.

The data described here supports a part of the research article "Effect of 4­HNE Modification on ZU5-ANK Domain and the Formation of Their Complex with ß­Spectrin: A Molecular Dynamics Simulation Study" [1]. Dataset on Gaff force field parameters of AMBER is provided for the non-standard arginine resulting of reaction with 4-hydroxy-2-nonenal (4-HNE), the major secondary product of lipids peroxidation. Arg-HNE 2-pentilpyrrole adduct is part of the 4-hydroxyalkenals described in various physiopathological disorders related to increased oxidative stress. Data include a framework for derivation of missing bonds, angles and dihedral parameters for modified arginine, alongside optimized partial charges derived with Restrained Electrostatic Potential (RESP) method and the new force field parameters obtained by quantum mechanicals methods (QM) using Hartree-Fock (HF)/6 - 31G** level of theory. Benchmark as a graphics tutorial summary steps to obtained new parameters and the validation of non-standard amino acids is presented. The new residue constructed is put available to the scientific community to perform molecular dynamics simulations of modified 4-HNE proteins on arginine residue and complete the set of data parameters for nucleophilic residues with this reactive aldehyde ADDIN EN.CITE ADDIN EN.CITE.DATA [2]. Data that could be used for the researchers interested in the role of protein oxidation as mediator in cellular pathophysiological.

Residues of Fluoroquinolone Antibiotics Induce Carbonylation and Reduce In Vitro Digestion of Sarcoplasmic and Myofibrillar Beef Proteins.

Although the impact of oxidation on human health has been of growing interest, the oxidation of proteins, major component of meat, has received little attention. This paper describes the in vitro effect of five fluoroquinolones (FQs) on carbonylation of sarcoplasmic and myofibrillar proteins of beef when found at concentrations close to the maximum residue limit (MRL). Samples were treated individually with the FQs, determining in each protein fraction the carbonyl index, protein content and oxidized proteins identification, using 2,4-dinitrophenyhydrazine (DNPH) alkaline assay, Western blot and Bradford methods, and mass spectrometry, respectively. Besides, the in vitro effect of these residues on gastric and duodenal digestion of proteins was evaluated. The carbonylation induced by FQs affected both protein fractions being significant with respect to the blank in 73.3% of cases. This damage was correlated with loss of solubility and digestibility, with sarcoplasmic proteins the most affected. Danofloxacin and enrofloxacin were the FQs with greatest oxidant effects, especially affecting glycolysis and glycogen proteins. Our results suggest that these residues induce irreversible oxidative damage on the main beef proteins and could affect their nutritional value.

Effect of 4-HNE Modification on ZU5-ANK Domain and the Formation of Their Complex with ß-Spectrin: A Molecular Dynamics Simulation Study.

4-HNE-modified ankyrins have been described in diseases such as diabetes, renal failure, G6PD deficient, sickle cell trait, and P. falciparum infected erythrocytes with different AB0 blood groups. However, effects at the atomic level of this carbonylation on structure and function of modified protein are not yet fully understood. We present a study based on molecular dynamics simulations of nine 4-HNE modified residues of the ZU5-ANK ankyrin domain with ß-spectrin and their binding energy profiles. Results show that 4-HNE induced local conformational changes over all protein systems evaluated, increased mobility in the modification sites, and localized structural changes between the positively charged patch of the ZU5-ANK domain. Carbonylation with 4-HNE on lysine residues decreased the affinity between ZU5-ANK and the 14-ß-spectrin repeat by reducing electrostatic and van der Waals interactions. The presented work provides further insight into understanding the loss of human erythrocyte deformation capacity under conditions of oxidative stress in different diseases.

Protein Carbonylation As a Biomarker of Heavy Metal, Cd and Pb, Damage in Paspalum fasciculatum Willd. ex Flüggé.

Heavy metal tolerant plants have phytoremediation potential for the recovery of contaminated soils, and the characterization of their metabolic adaptation processes is an important starting point to elucidate their tolerance mechanisms at molecular, biochemical and physiological levels. In this research, the effects of Cd and Pb on growth and protein carbonylation in tissues of Paspalum fasciculatum exposed to 30 and 50 mg·Kg-1 Cd and Pb respectively were determined. P. fasciculatum seedlings exposed to metals grew more than controls until 60 days of cultivation and limited their oxidative effects to a reduced protein group. Carbonyl indexes in leaf and root proteins reached a significant increase concerning their controls in plants exposed 30 days to Cd and 60 days to Pb. From the combined approach of Western Blot with Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis (SDS-PAGE) and protein analysis by Matrix Asisted Laser Desorption/Ionisation - Time Of Flight (MALDI-TOF/TOF) mass spectrometry, chloroplastic proteins were identified into the main oxidative stress-inducible proteins to Cd and Pb, such as subunits α, γ of ATP synthetase, Chlorophyll CP26 binding protein, fructose-bisphosphate aldolase and long-chain ribulose bisphosphate carboxylase (RuBisCO LSU). Cd generated damage in the photosynthetic machinery of the leaves of P. fasciculatum into the first 30 days of treatment; five of the oxidized proteins are involved in photosynthesis processes. Moreover, there was a proteolytic fragmentation of the RuBisCO LSU. Results showed that intrinsic tolerance of P. fasciculatum to these metals reached 60 days in our conditions, along with the bioaccumulating appreciable quantities of metals in their roots.

Sickle Cell Trait Induces Oxidative Damage on Plasmodium falciparum Proteome at Erythrocyte Stages.

The presence of hemoglobin A-S (HbAS) in erythrocytes has been related to the high production of reactive oxygen species (ROS) and an increased in intracellular oxidative stress that affects the progress of Plasmodium erythrocytic cycle life and attenuates its serious clinical symptoms. Nevertheless, oxidative effects on P. falciparum proteome across the intraerythrocytic cycle in the presence of HbAS traits have not been described yet. Here, an immune dot-blot assay was used to quantify the carbonyl index (C.I) on P. falciparum 3D7 proteome at the different asexual erythrocytic stages. Protein carbonylation on parasites cultivated in erythrocytes from two donors with HbAS increased 5.34 ± 1.42 folds at the ring stage compared to control grown in hemoglobin A-A (HbAA) red blood cells. Whereas at trophozoites and schizonts stages were augmented 2.80 ± 0.52 and 3.05 ± 0.75 folds, respectively. Besides proteins involved in processes of the stress response, recognition and invasion were identified from schizonts carbonylated bands by combining SDS-PAGE with MALDI-TOF-TOF analysis. Our results reinforce the hypothesis that such oxidative modifications do not appear to happen randomly, and the sickle cell trait affects mainly a small fraction of parasite proteins particularly sensitive to ROS.

Quantification of microplastics along the Caribbean Coastline of Colombia: Pollution profile and biological effects on Caenorhabditis elegans.

The Caribbean Coast of Colombia has a flourishing plastic industry with weak and insufficient waste management policies and practices, leading to plastic pollution along its touristic beaches. In this work, primary and secondary microplastics (MPs) were surveyed at four different locations along the Colombian Caribbean Coast. Primary microplastics, specifically white new plastic pellets, represented the largest amount of MPs found, with densities decreasing in the order Cartagena > Coveñas > Puerto Colombia > Riohacha. This distribution was connected to the vicinity of MPs sources, marine currents and wind direction. The presence of secondary MPs was associated with urban centers and proximity to river mouths. The FTIR characterization showed polyethylene as the predominant resin type, with different degrees of surface oxidation. Aqueous extracts from sampled MPs were tested on Caenorhabditis elegans. Secondary MPs elicited greater toxicological responses than pellets, especially those from Cartagena Bay, suggesting MPs act as carriers for biologically-active pollutants.