http://repositorio.ufc.br/handle/riufc/364 Sat, 15 Aug 2026 21:40:49 GMT 2026-08-15T21:40:49Z http://repositorio.ufc.br/handle/riufc/87488 Título: Purificação, caracterização e avaliação da atividade anti-helmíntica de uma peroxidase do látex de Calotropis procera (Ait.)R. Br Autor(es): Bezerra, Leandro de Paula Abstract: Latex is a complex fluid enriched in hydrolytic enzymes, such as proteases and chitinases; however, the occurrence and role of peroxidases in this material remain poorly understood. Peroxidases (EC 1.11.1.7) are widely distributed across biological systems due to their involvement in numerous physiological processes. These enzymes are classified as heme and non-heme peroxidases, with heme-containing peroxidases accounting for approximately 84% of all described peroxidases, according to the RedOxiBase database. Among them, class III plant peroxidases constitute a large multigene family of heme-dependent enzymes that share a similar three-dimensional structure and a common catalytic mechanism for hydrogen peroxide degradation. They are involved in the metabolism of reactive oxygen species (ROS), hormone synthesis and degradation, fruit growth, defense responses, and cell wall synthesis and maintenance. In this context, we report the purification and structural characterization of an atypical peroxidase (CpLP-POX) isolated from the latex of C. procera. Initial analyses revealed a high-molecular-weight enzyme (~112 kDa) that dissociated into ~35 kDa subunits under denaturing conditions, suggesting a trimeric organization. Structural modeling corroborated the homotrimeric assembly of the protein. The CpLP-POX monomers exhibited a conserved catalytic triad (Arg–His–Asn), four disulfide bonds, and the overall fold characteristic of class III plant peroxidases. The enzyme displayed maximal activity at pH 6.0 and 40 °C and was inhibited by EDTA, sodium azide, and thiol-reducing agents, consistent with the profile of Ca2+-associated heme-dependent peroxidases. CpLP-POX did not exhibit inhibitory effects on nematode egg hatching, suggesting that its physiological role may be more closely related to oxidative defense or protection against microbial invasion. Collectively, these findings provide the first biochemical and structural evidence of a trimeric peroxidase in plants, expanding the known structural diversity of this enzyme class and contributing to a deeper understanding of its biological functions. Tipo: Tese Thu, 01 Jan 2026 00:00:00 GMT http://repositorio.ufc.br/handle/riufc/87488 2026-01-01T00:00:00Z http://repositorio.ufc.br/handle/riufc/87393 Título: Water deficit tolerance in cowpea plants is associated with coordinated metabolic and physiological mechanisms involving roots and leaves Autor(es): León, Danny Alexander Camargo Abstract: Drought reduces the productivity of agricultural crops and restricts the use of extensive areas for agricultural production. Several studies have reported that cowpea exhibits a variety of physiological, biochemical, and phenotypic adaptations that enable it to survive and maintain productivity under drought conditions, making it a suitable model for studies associated with plant responses to water deficit. The most commonly reported acclimatizations include the regulation of stomatal conductance, the accumulation of compatible osmolytes, and adjustments in leaf morphology. On the other hand, the underlying mechanisms that allow cowpea to modify root system architecture under water deficit conditions have been poorly studied. Therefore, in the present thesis, through an integrated approach, we investigated water deficit tolerance mechanisms in cowpea plants, emphasizing the importance of metabolic and physiological responses. We delved into the interactions between the plant's capacity to adjust root growth and changes in the metabolic profile, as well as other biochemical and physiological adjustments under water deficit conditions. For this purpose, 21-day-old cowpea plants were subjected to progressive irrigation withdrawal, resulting in three water regimes: a well-watered control (75% of substrate water-holding capacity), moderate water deficit (25% of substrate water-holding capacity), and severe water deficit (0% of substrate water-holding capacity) for seven days. After five days of water withdrawal, moderately stressed plants were rewatered and allowed to recover to control conditions for two consecutive days. Leaf and root samples were collected for GC-MS-based metabolomic analyses, including amino acids, soluble sugars, and organic acids. In addition, the activities of enzymes related to nitrogen metabolism, including glutamine synthetase, glutamate synthase, and glutamate dehydrogenase, were determined. Physiological analyses were also performed, including gas exchange, osmotic adjustment capacity, root and shoot biomass, and indicators of water deficit, such as relative water content, membrane damage, and water potential. The results revealed progressive reductions in photosynthesis, stomatal conductance, and water potential under increasing levels of water deficit, accompanied by contrasting responses between roots and shoots. Whereas shoot growth was markedly reduced, roots exhibited increased biomass under moderate water deficit, suggesting a redistribution of resources to enhance water acquisition. Following rehydration, root biomass decreased and shoot growth resumed, indicating a functional reorganization of the root system during recovery. Enzymatic responses, particularly those involving glutamine synthetase and glutamate dehydrogenase, indicated a coordinated reorganization of nitrogen metabolism in leaves and roots under water deficit conditions. In parallel, plants exhibited osmotic adjustment in both organs, associated with the accumulation of amino acids, soluble sugars, and glycine betaine. Although proline accumulated substantially during stress, its relative contribution to osmotic adjustment was limited, indicating that maintenance of tissue turgor depends on the coordinated action of multiple osmolytes. Integrated physiological and metabolomic analyses revealed strong associations among plant water status, nitrogen metabolism, amino acid accumulation, and growth- related traits. Consistent alterations in 2-oxoglutarate and other organic acids suggest the involvement of these metabolites in the integration of carbon and nitrogen metabolism during water deficit. Furthermore, the high recovery capacity observed for several physiological and biochemical traits demonstrates that drought tolerance in cowpea depends on coordinated responses between roots and leaves involving physiological, biochemical, and metabolic adjustments that operate in an integrated manner during both stress and recovery. Tipo: Tese Mon, 01 Jan 2024 00:00:00 GMT http://repositorio.ufc.br/handle/riufc/87393 2024-01-01T00:00:00Z http://repositorio.ufc.br/handle/riufc/87204 Título: Caracterização estrutural e propriedades anti-inflamatória e antiviral de polissacarídeos sulfatados de algas marinhas do genêro Hypnea Autor(es): Barros, Francisco Clark Nogueira Abstract: Marine seaweeds fave demonstrated to be a importante source of bioactive molecules. Algae from the genus Hypnea are known for the production carrageenan polysaccharides, a hidrocolloid used in pharmaceutical, cosmetic, food and biotechnological industries. Due to the morphological simplicity of these organisms, the identification of these organisms based on anatomic and morphological characters has led to mistakes. To overcome this problem, several studies have proposed the use of gene markers to aid in the identification of these organisms. In this work, specimens of the species Hypnea pseudomusciformis variety. “musciformis” “and “nigrescens” and Hypnea cervicornis, collected on the coast of Ceará, São Paulo and Bahia were studied. The identification of H. pseudomusciformis was made through morphologial characters. Though, for H. cervicornis a morphological analysis followed by the sequence of two gene markers (COI-5P and rbcL) were used. Later, sulfated polysaccharides were isolated from each group. The chemical structure of these polymers was characterized by elemental microanalysis, FT-IT, NMR and High Performance Size Exclusion Chromatography. The polysaccharide isolated from H. pseudomusciformis var. “musciformis” (HmP) showed a degree of substitution for sulfate groups (DS sulfate) of 0.88, while the variety nigrescens presented a DS sulfate = 1.05. The polysaccharide from H. cervicornis (HcP) had a DS sulfate of 0.79. The peak molar mass of HmP was 123.6 kDa. To HnP, the value observed was 71.4 kDa, while HcP peak molar mass was estimated in 70.2 kDa. FT-IR spectra of HmP, HnP and HcP revealed the presence of bands with similiar wavenumbers, characteristic of kappa-carrageenan. NMR spectra of HmP and HnP revealed that 7.3% of HmP and 9.2% of HnP were ι-carrageenan. NMR also demonstrated that 95% HcP had 95% of kappa-carrageenan and only 5% as ι-carrageenan. HcP was also evaluated regarding its effect on classic models of inflammation in mice. HcP (30 mg/kg; i.p.) reduced paw edema induced by carrageenin in 69% and the migration of inflammatory cells, as seen by mieloperoxidase (MPO) counting reduction of 53%. HcP also reduced the migration of neutrophils to the peritoneal cavity induced by carrageenin and fMLP. Though, HcP did not reduced the osmotic/vascular inflammatory response induced by histamine or 48/80 compound. Yet, HcP (30 mg/kg; i.p.) exerted antinociceptive effect in mice when inflammation was induced by formalin injection in the animal paw. The licking time was significantly reduced in the second phase, which is known to involve inflammatoy response. Though, no effect was detected in the first phase, regarded to be neurogenic. The analgesic effect of HcP was confirmed by a hyperssensitivity test (Von Frey assay). The polysaccharides isolated from H. pseudomusciformis were evaluated regarding their potential for neutralizing virus, especially HIV. In a first step it was verified the bond capacity of HnP and HmP against several viral proteins by ELISA. Both polysaccharides bound to te glycoproteins gp120/gp140 from HIV, S2-rGP, S2-GPe and ZEBOV rGP from Ebola virus, Dengue III, Chikungunya E1 and E2. Then, kinetics and affinity of HnP binding to the glyprotein gp140 from HIV was evaluated by Surface Plasmon Resonance in a gold CM-5 chip. The affinity constanto (KD =1,450 x 10-9) observed for HnP binding to gp120 was determined. Later, it was analysed HnP neutralizing effect against HIV cultivated in Tzmbl cells expressing luciferase. It was tested one virus of clade B in a live form called HIV Bal (IC50 = 73,51 μg/mL; IC90 = 661,59 μg/mL) and the four HIV pseudoviruses of clades B and C as follows: Bal pseudovirus (IC50 = 54,84 μg/mL; IC90 = 493,56 μg/mL), PCAAN S342 (IC50 = 36,67 μg/mL; IC90 = 330,03 μg/mL), SC422-661.8 (IC50 = 25,79 μg/mL; IC90 = 232,11 μg/mL) and ZM53M.PB12 SVPC11 (IC50 = 70,85 μg/mL; IC90 = 637,65 μg/mL). In addition, HnP and HmP effect on Rabies virus cultivated in BHK21 cells. HnP neutralized Rabies CVS11 pseudovirus with IC50 = 71,04 μg/mL and IC90 = 639,36 μg/mL. Though, HmP effect against Rabies CVS11 pseudovirus was significantly higher (IC50 = 10,99 μg/mL; IC90 = 98,91 μg/mL). It was also evaluated HnP and HmP cytotoxicity against the two cell lines used for viral cultivation by the Lactate dehydrogenase assay (LDH). No HnP toxicity was found against Tzmbl and BHK21 cells until 375 μg/mL when incubated for 2h and until 750 μg/mL when incubated for 9h. Similar result was found was found to Tzmbl and BHK21 cells incubated with HmP. Finally, it is possible to infer that HcP revealed and important anti-inflammatory potential and that HnP and HmP demonstrated a potent antiviral effect and could be used in the development of new therapeutic approaches. Tipo: Tese Mon, 01 Jan 2018 00:00:00 GMT http://repositorio.ufc.br/handle/riufc/87204 2018-01-01T00:00:00Z http://repositorio.ufc.br/handle/riufc/86861 Título: Inibidores de proteases de plantas como modelos estruturais para o desenho de peptídeos bioinspirados com potencial uso clínico sobre as proteases principais de SARS-COV-2, SARS-COV-1 e MERS-COV Autor(es): Lima, Adrianne Maia Abstract: Human coronaviruses (HCoVs) associated with severe infections, such as MERS-CoV, SARS- CoV-1, and SARS-CoV-2, remain a global threat due to limited effective antivirals, reduced efficacy of vaccines against new variants, and the risk of zoonotic outbreaks. The main viral protease (Mpro), essential for replication and highly conserved among HCoVs, represents a key therapeutic target. Given the potential of plant protease inhibitors (PIs) to modulate viral enzymes, this study aimed to identify plant-derived PIs targeting Mpro and evaluate their use as scaffolds for bioinspired antiviral peptides. A structural database of plant PIs was compiled from Protein Data Bank (PDB) entries, focusing on cysteine protease inhibitors. Molecular docking (FRODOCK 3.12 and ClusPro 2.0), molecular dynamics simulations (100 ns, GROMACS 2018.4), and quantum mechanical calculations (DMol3) revealed that phytocystatins from Ananas comosus and Sesamum indicum interact with allosteric sites of Mpro, with interaction energies of -225.7 and -301.9 kcal/mol, respectively. CASTp server analyses indicated that these interactions caused significant reductions in the active site area and volume of Mpro. Based on the sequences of these phytocystatins, two bioinspired peptides were designed — KFD-Pep1 (1276.31 Da) and MLL-Pep2 (1294.69 Da) — showing favorable in silico properties, including predicted cell penetration, positive net charges (+3 and +4), hydrophobicity (50% and 40%), Boman indices (2.1 and 1.34 kcal/mol), α-helical structures, simulated intestinal stability, and predictions of being non-hemolytic and non-allergenic. Notably, MLL-Pep2 also displayed an amphipathic nature. Molecular docking analyses showed that both peptides interact with conserved catalytic regions of Mpro of SARS-CoV-2 (wild- type, naturally occurring mutants, and the E166R-induced mutant), SARS-CoV-1, and MERS- CoV. ADMET profiling predicted favorable pharmacokinetic properties and low toxicity, comparable to the clinically approved inhibitor nirmatrelvir. These findings highlight KFD- Pep1 and MLL-Pep2 as promising candidates for further studies and support plant protease inhibitors as valuable sources for broad-spectrum antiviral development against HCoVs. Tipo: Tese Wed, 01 Jan 2025 00:00:00 GMT http://repositorio.ufc.br/handle/riufc/86861 2025-01-01T00:00:00Z