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Multidrug-resistant Cryptococcus neoformans is an encapsulated yeast causing a high mortality rate in immunocompromised patients. Recently, the synthetic peptide Mo-CBP3-PepII emerged as a potent anticryptococcal molecule with an MIC50 at low concentration. Here, the mechanisms of action of Mo-CBP3-PepII were deeply analyzed to provide new information about how it led C. neoformans cells to death. Light and fluorescence microscopies, analysis of enzymatic activities, and proteomic analysis were employed to understand the effect of Mo-CBP3-PepII on C. neoformans cells. Light and fluorescence microscopies revealed Mo-CBP3-PepII induced the accumulation of anion superoxide and hydrogen peroxide in C. neoformans cells, in addition to a reduction in the activity of superoxide dismutase (SOD), ascorbate peroxidase (APX), and catalase (CAT) in the cells treated with Mo-CBP3-PepII. In the presence of ascorbic acid (AsA), no reactive oxygen species (ROS) were detected, and Mo-CBP3-PepII lost the inhibitory activity against C. neoformans. However, Mo-CBP3-PepII inhibited the activity of lactate dehydrogenase (LDH) ergosterol biosynthesis and induced the decoupling of cytochrome c (Cyt c) from the mitochondrial membrane. Proteomic analysis revealed a reduction in the abundance of proteins related to energetic metabolism, DNA and RNA metabolism, pathogenicity, protein metabolism, cytoskeleton, and cell wall organization and division. Our findings indicated that Mo-CBP3-PepII might have multiple mechanisms of action against C. neoformans cells, mitigating the development of resistance and thus being a potent molecule to be employed in the production of new drugs against C. neoformans infections.
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Staphylococcus aureus is a human pathogen known to be resistant to antibiotics since the mid-20th century and is constantly associated with hospital-acquired infections. S. aureus forms biofilms, which are complex surface-attached communities of bacteria held together by a self-produced polymer matrix consisting of proteins, extracellular DNA, and polysaccharides. Biofilms are resistance structures responsible for increasing bacterial resistance to drugs by 1000 times more than the planktonic lifestyle. Therefore, studies have been conducted to discover novel antibacterial molecules to prevent biofilm formation and/or degrade preformed biofilms. Synthetic antimicrobial peptides (SAMPs) have appeared as promising alternative agents to overcome increasing antibiotic resistance. Here, the antibiofilm activity of eight SAMPs, in combination with the antibiotic ciprofloxacin, was investigated in vitro. Biofilm formation by S. aureus was best inhibited (76%) by the combination of Mo-CBP3-PepIII (6.2 µg mL-1) and ciprofloxacin (0.39 µg mL-1). In contrast, the highest reduction (60%) of the preformed biofilm mass was achieved with RcAlb-PepII (1.56 µg mL-1) and ciprofloxacin (0.78 µg mL-1). Fluorescence microscopy analysis reinforced these results. These active peptides formed pores in the cellular membrane of S. aureus, which may be related to the enhanced ciprofloxacin's antibacterial activity. Our findings indicated that these peptides may act with ciprofloxacin and are powerful co-adjuvant agents for the treatment of S. aureus infections.
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BACKGROUND: Chitinases are plant defense-related proteins with a high biotechnological potential to be applied in agriculture. OBJECTIVES: This study aimed to purify a chitinase from the latex of Ficus benjamina. METHODS: An antifungal class I chitinase, named FbLx-Chi-1, was purified from the latex of Ficus benjamina after precipitation with 30-60% ammonium sulfate and affinity chromatography on a chitin column and antifungal potential assay against phytopathogenic fungi important to agriculture. RESULTS: FbLx-Chi-1 has 30 kDa molecular mass, as estimated by SDS-PAGE and the optimal pH and temperature for full chitinolytic activity were 5.5 and 60ºC, respectively. FbLx-Chi-1 is a high pH-, ion-tolerant and thermostable protein. Importantly, FbLx-Chi-1 hindered the growth of the phytopathogenic fungi Colletotrichum gloeosporioides, Fusarium pallidoroseum, and Fusarium oxysporum. The action mode of FbLx-Chi-1 to hamper F. pallidoroseum growth seems to be correlated with alterations in the morphology of the hyphal cell wall, increased plasma membrane permeability, and overproduction of reactive oxygen species. CONCLUSION: These findings highlight the biotechnological potential of FbLx-Chi-1 to control important phytopathogenic fungi in agriculture. In addition, FbLx-Chi-1 could be further explored to be used in industrial processes such as the large-scale environmentally friendly enzymatic hydrolysis of chitin to produce its monomer N-acetyl-ß-D-glucosamine, which is employed for bioethanol production, in cosmetics, in medicine, and for other multiple applications.
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Quitinases , Ficus , Antifúngicos/farmacologia , Antifúngicos/química , Látex , Ficus/metabolismo , Espécies Reativas de Oxigênio , Quitinases/farmacologia , Quitinases/química , Quitinases/metabolismo , Quitina/farmacologia , Quitina/química , Parede Celular/metabolismo , Membrana Celular/metabolismoRESUMO
L: operculata is a plant commonly found in the North and Northeast of Brazil. Although the regional population knows its medicinal potential, there are few scientific studies about its antimicrobial potential. Thus, this study aimed to characterize the proteins from L. operculata seeds extracted using different solutions and evaluate their antimicrobial potentials. The protein extracts obtained with NaCl and sodium acetate buffer presented the best inhibitory activities against Candida albicans and C. krusei. The study of the mechanism of action revealed proteins from L. operculata seeds induced pore formation on the membrane and ROS overaccumulation. Scanning Electron Microscopy images also showed severe morphological changes in Candida albicans and C. krusei. Proteins from L.operculata seeds did not show antibacterial activity. The enzymatic assays revealed the presence of proteolytic enzymes, serine and cysteine protease inhibitors, and chitinases in both protein extracts. Proteomic analysis by LC-ESI-MS/MS identified 57 proteins related to many biological processes, such as defense to (a)biotic stress, energetic metabolism, protein folding, and nucleotide metabolism. In conclusion, the L. operculata seed proteins have biotechnological potential against the human pathogenic yeasts Candida albicans and C. krusei.
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Candida albicans , Luffa , Antibacterianos , Humanos , Testes de Sensibilidade Microbiana , Proteômica , Sementes , Espectrometria de Massas em TandemRESUMO
Cryptococcus neoformans is a human-pathogenic yeast responsible for pneumonia and meningitis, mainly in patients immunocompromised. Infections caused by C. neoformans are a global health concern. Synthetic antimicrobial peptides (SAMPs) have emerged as alternative molecules to cope with fungal infections, including C. neoformans. Here, eight SAMPs were tested regarding their antifungal potential against C. neoformans and had their mechanisms of action elucidated by fluorescence and scanning electron microscopies. Five SAMPs showed an inhibitory effect (MIC50) on C. neoformans growth at low concentrations. Fluorescence microscope (FM) revealed that SAMPs induced 6-kDa pores in the C. neoformans membrane. Inhibitory assays in the presence of ergosterol revealed that some peptides lost their activity, suggesting interaction with it. Furthermore, FM analysis revealed that SAMPs induced caspase 3/7-mediated apoptosis and DNA degradation in C. neoformans cells. Scanning Electron Microscopy (SEM) analysis revealed that peptides induced many morphological alterations such as cell membrane, wall damage, and loss of internal content on C. neoformans cells. Our results strongly suggest synthetic peptides are potential alternative molecules to control C. neoformans growth and treat the cryptococcal infection.
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The outbreak caused by SARS-CoV-2 has taken many lives worldwide. Although vaccination has started, the development of drugs to either alleviate or abolish symptoms of COVID-19 is still necessary. Here, four synthetic peptides were assayed regarding their ability to protect Vero E6 cells from SARS-CoV-2 infection and their toxicity to human cells and zebrafish embryos. All peptides had some ability to protect cells from infection by SARS-CoV-2 with the D614G mutation. Molecular docking predicted the ability of all peptides to interact with and induce conformational alterations in the spike protein containing the D614G mutation. PepKAA was the most effective peptide, by having the highest docking score regarding the spike protein and reducing the SARS-CoV-2 plaque number by 50% (EC50) at a concentration of 0.15 mg mL-1. Additionally, all peptides had no toxicity to three lines of human cells as well as to zebrafish larvae and embryos. Thus, these peptides have potential activity against SARS-CoV-2, making them promising to develop new drugs to inhibit cell infection by SARS-CoV-2.
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Antimicrobial resistance has been increasing globally, posing a global public health risk. It has prompted the scientific community to look for alternatives to traditional drugs. Antimicrobial Peptides (AMPs) have stood out in this context because they have the potential to control infectious diseases while causing no or little harm to mammalian cells. In the present study, three peptides, JcTI-PepI, JcTI-PepII, and JcTI-PepIII, were designed and tested for antimicrobial activity based on the primary sequence of JcTI-I, a 2S albumin with trypsin inhibitory activity from Jatropha curcas. JcTI-PepI strongly inhibited C. krusei growth, and it caused severe disruptions in cellular processes and cell morphology. C. krusei cells treated with JcTI-PepI showed indicative of membrane permeabilization and overproduction of Reactive Oxygen Species. Moreover, the yeast's ability to acidify the medium was severely compromised. JcTI-PepI was also effective against pre-formed biofilm and did not harm human erythrocytes and Vero cells. Overall, these characteristics indicate that JcTI-PepI is both safe and effective against C. krusei, an intrinsically resistant strain that causes serious health problems and is frequently overlooked. It implies that this peptide has a high potential for use as a new antimicrobial agent in the future.
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Anti-Infecciosos , Jatropha , Animais , Anti-Infecciosos/farmacologia , Chlorocebus aethiops , Humanos , Mamíferos , Testes de Sensibilidade Microbiana , Peptídeos/farmacologia , Inibidores da Tripsina , Células VeroRESUMO
C. albicans and C. parapsilosis are biofilm-forming yeasts responsible for bloodstream infections that can cause death. Synthetic antimicrobial peptides (SAMPs) are considered to be new weapons to combat these infections, alone or combined with drugs. Here, two SAMPs, called Mo-CBP3-PepI and Mo-CBP3-PepIII, were tested alone or combined with nystatin (NYS) and itraconazole (ITR) against C. albicans and C. parapsilosis biofilms. Furthermore, the mechanism of antibiofilm activity was evaluated by fluorescence and scanning electron microscopies. When combined with SAMPs, the results revealed a 2- to 4-fold improvement of NYS and ITR antibiofilm activity. Microscopic analyses showed cell membrane and wall damage and ROS overproduction, which caused leakage of internal content and cell death. Taken together, these results suggest the potential of Mo-CBP3-PepI and Mo-CBP3-PepIII as new drugs and adjuvants to increase the activity of conventional drugs for the treatment of clinical infections caused by C. albicans and C. parapsilosis.
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The recent outbreak caused by SARS-CoV-2 continues to threat and take many lives all over the world. The lack of an efficient pharmacological treatments are serious problems to be faced by scientists and medical staffs worldwide. In this work, an in silico approach based on the combination of molecular docking, dynamics simulations, and quantum biochemistry revealed that the synthetic peptides RcAlb-PepI, PepGAT, and PepKAA, strongly interact with the main protease (Mpro) a pivotal protein for SARS-CoV-2 replication. Although not binding to the proteolytic site of SARS-CoV-2 Mpro, RcAlb-PepI, PepGAT, and PepKAA interact with other protein domain and allosterically altered the protease topology. Indeed, such peptide-SARS-CoV-2 Mpro complexes provoked dramatic alterations in the three-dimensional structure of Mpro leading to area and volume shrinkage of the proteolytic site, which could affect the protease activity and thus the virus replication. Based on these findings, it is suggested that RcAlb-PepI, PepGAT, and PepKAA could interfere with SARS-CoV-2 Mpro role in vivo. Also, unlike other antiviral drugs, these peptides have no toxicity to human cells. This pioneering in silico investigation opens up opportunity for further in vivo research on these peptides, towards discovering new drugs and entirely new perspectives to treat COVID-19.Communicated by Ramaswamy H. Sarma.
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COVID-19 , SARS-CoV-2 , Humanos , Domínio Catalítico , Simulação de Acoplamento Molecular , Peptídeos/farmacologia , Peptídeo Hidrolases , Inibidores de Proteases/farmacologia , Simulação de Dinâmica MolecularRESUMO
Vaccines could be the solution to the current SARS-CoV-2 outbreak. However, some studies have shown that the immunological memory only lasts three months. Thus, it is imperative to develop pharmacological treatments to cope with COVID-19. Here, the in silico approach by molecular docking, dynamic simulations and quantum biochemistry revealed that ACE2-derived peptides strongly interact with the SARS-CoV-2 RBD domain of spike glycoprotein (S-RBD). ACE2-Dev-PepI, ACE2-Dev-PepII, ACE2-Dev-PepIII and ACE2-Dev-PepIV complexed with S-RBD provoked alterations in the 3D structure of S-RBD, leading to disruption of the correct interaction with the ACE2 receptor, a pivotal step for SARS-CoV-2 infection. This wrong interaction between S-RBD and ACE2 could inhibit the entry of SARS-CoV-2 in cells, and thus virus replication and the establishment of COVID-19 disease. Therefore, we suggest that ACE2-derived peptides can interfere with recognition of ACE2 in human cells by SARS-CoV-2 in vivo. Bioinformatic prediction showed that these peptides have no toxicity or allergenic potential. By using ACE2-derived peptides against SARS-CoV-2, this study points to opportunities for further in vivo research on these peptides, seeking to discover new drugs and entirely new perspectives to treat COVID-19.Communicated by Ramaswamy H. Sarma.
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Tratamento Farmacológico da COVID-19 , Glicoproteína da Espícula de Coronavírus , Enzima de Conversão de Angiotensina 2 , Humanos , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Peptídeos/química , Peptídeos/farmacologia , Peptidil Dipeptidase A/química , Ligação Proteica , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/metabolismoRESUMO
Fungal contamination is among the main reasons for food spoilage, affecting food safety and the economy. Among fungi, Penicillium digitatum is a major agent of this problem. Here, the in vitro activity of eight synthetic antimicrobial peptides was assessed against P. digitatum, and their action mechanisms were evaluated. All peptides were able to inhibit fungal growth. Furthermore, atomic force and fluorescence microscopies revealed that all peptides targeted the fungal membrane leading to pore formation, loss of internal content, and death. The induction of high levels of reactive oxygen species (ROS) was also a mechanism employed by some peptides. Interestingly, only three peptides (PepGAT, PepKAA, and Mo-CBP3-PepI) effectively control P. digitatum colonization in orange fruits, at a concentration (50 µg mL-1) 20-fold lower than the commercial food preservative (sodium propionate). Altogether, PepGAT, PepKAA, and Mo-CBP3-PepI showed high biotechnological potential as new food preservatives to control food infection by P. digitatum.
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Citrus sinensis , Penicillium , Frutas , Proteínas Citotóxicas Formadoras de PorosRESUMO
Resistant nematodes are not affected by the most common drugs commercially available. In the search for new anthelmintics, peptides have been investigated. Here, a linear synthetic peptide named RcAlb-PepIII bioinspired from the antimicrobial protein Rc-2S-Alb was designed, synthesized, and tested against barber pole worm Haemonchus contortus. The physicochemical properties of the peptide, the 3D structure model, the egg hatch inhibition, and larval development inhibition of H. contortus were carried out. Additionally, the ultrastructure of the nematode after treatment with the peptide was evaluated by atomic force microscopy. The RcAlb-PepIII inhibited the larval development of H. contortus with an EC50 of 90 µM and did not affect egg hatch. Atomic force microscopy reveals the high affinity of RcAlb-PepIII with the cuticle of H. contortus in the L2 stage. It also shows the deposition of RcAlb-PepIII onto the surface of the cuticle, forming a structure similar to a film that reduces the roughness and mean square roughness (Rq) of it. In conclusion, the bioinspired RcAlb-PepIII has the potential to be used as a new anthelmintic compound to control gastrointestinal nematode parasites.
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Recently, the dramatic emergence of antimicrobial resistance has received attention from World Health Organization. Synthetic antimicrobial peptides (SAMPs) are considered new weapons to fight against infections caused by multi-drug resistant pathogens. Here, the authors provide an overview of the current research on SAMPs. The focus is SAMPs, how to design them, which features must be considered during design, and comparison with natural peptides. This review also includes a discussion about the natural AMPs, mechanisms of action and applications as new drugs or even as adjuvants molecules to enhance commercial drugs activity. The advances in chemical synthesis have reduced the cost to produce synthetic peptides open ways to achieve new antimicrobial agents. Therefore, synthetic peptides are new promising molecules to safeguard human and animal health.
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Anti-Infecciosos/química , Anti-Infecciosos/farmacologia , Desenho de Fármacos , Resistência Microbiana a Medicamentos , Proteínas Citotóxicas Formadoras de Poros/química , Proteínas Citotóxicas Formadoras de Poros/farmacologia , Animais , Anti-Infecciosos/síntese química , Bactérias/efeitos dos fármacos , Infecções Bacterianas/tratamento farmacológico , Técnicas de Química Sintética , Resistência Microbiana a Medicamentos/efeitos dos fármacos , Resistência a Múltiplos Medicamentos/efeitos dos fármacos , Fungos/efeitos dos fármacos , Humanos , Micoses/tratamento farmacológico , Proteínas Citotóxicas Formadoras de Poros/síntese químicaRESUMO
AIMS: According to the WHO, 20-25% of people worldwide are affected by skin infections caused by dermatophytes, such as those of the Trichophyton genus. Additionally, several dermatophytes have developed resistance to drugs such as griseofulvin and itraconazole. This study tested 2S albumins-derived antimicrobial peptides (AMPs) as alternative antidermatophytic molecules. MAIN METHODS: Membrane pore formation assays, tests to detect overproduction of ROS, scanning electron microscopy (SEM) and fluorescence microscopy (FM) were carried out to provide insight into the mechanisms of antidermatophytic action. KEY FINDINGS: All AMPs (at 50 µg mL-1) tested reduced the mycelial growth of T. mentagrophytes and T. rubrum by up to 95%. In contrast, using a concentration 20-fold higher, griseofulvin only inhibited T. mentagrophytes by 35%, while itraconazole was not active against both dermatophytes. Scanning electron and fluorescence microscopies revealed that the six AMPs caused severe damage to hyphal morphology by inducing cell wall rupture, hyphal content leakage, and death. Peptides also induced membrane pore formation and oxidative stress by overproduction of ROS. Based on the stronger activity of peptides than the commercial drugs and the mechanism of action, all six peptides have the potential to be either employed as models to develop new antidermatophytic drugs or as adjuvants to existing ones. SIGNIFICANCE: The synthetic peptides are more efficient than conventional drug to treat infection caused by dermatophytes being potential molecules to develop new drugs.
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Antifúngicos/farmacologia , Arthrodermataceae/efeitos dos fármacos , Griseofulvina/farmacologia , Itraconazol/farmacologia , Fragmentos de Peptídeos/farmacologia , Antifúngicos/síntese química , Arthrodermataceae/fisiologia , Técnicas de Química Sintética , Griseofulvina/síntese química , Humanos , Itraconazol/síntese química , Fragmentos de Peptídeos/síntese químicaRESUMO
BACKGROUND: Dermatophytes belonging to the Trichophyton genus are important human pathogens, but they have developed resistance to griseofulvin, the most common antifungal drug used to treat dermatophytosis. OBJECTIVE: This study was aimed to evaluate the antidermatophytic activity of synthetic peptides, as well as mechanisms of action and synergistic effect with griseofulvin. METHODS: Scanning electron microscopy (SEM), atomic force microscopy (AFM) and fluorescence microscopy (FM) were employed to understand the activity and the mechanism of action of peptides. RESULTS: Here we report that synthetic peptides at 50 µg/mL, a concentration 20-fold lower than griseofulvin, reduced the microconidia viability of T. mentagrophytes and T. rubrum by 100%, whereas griseofulvin decreased their viability by only 50% and 0%, respectively. The action mechanism of peptides involved cell wall damage, membrane pore formation and loss of cytoplasmic content. Peptides also induced overproduction of reactive oxygen species (ROS) and enhanced the activity of griseofulvin 10-fold against both fungi, suggesting synergistic effects, and eliminated the toxicity of this drug to human erythrocytes. Docking analysis revealed ionic and hydrophobic interactions between peptides and griseofulvin, which may explain the decline of griseofulvin toxicity when mixed with peptides. CONCLUSION: Therefore, our results strongly suggest six peptides with high potential to be employed alone as new drugs or as adjuvants to enhance the activity and decrease the toxicity of griseofulvin.
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Antifúngicos/farmacologia , Griseofulvina/farmacologia , Peptídeos/síntese química , Peptídeos/farmacologia , Esporos Fúngicos/efeitos dos fármacos , Trichophyton/efeitos dos fármacos , Descoberta de Drogas , Farmacorresistência Fúngica , Sinergismo Farmacológico , Humanos , Testes de Sensibilidade MicrobianaRESUMO
Lectins are a group of widely distributed and structurally heterogeneous proteins of nonimmune origin. These proteins have the ability to interact with glycans present on cell surfaces and elicit diverse biological activities. Machaerium acutifolium lectin (MaL) is an N-acetyl-D-glucosamine-binding lectin that exhibits antinociceptive activity via transient receptor potential cation channel subfamily V member 1 (TRPV1). Lectins that have the ability to recognize and interact with N-acetyl-D-glucosamine residues are potential candidates for studies of fungicidal activity. In this work, we show that MaL has antifungal activity against Candida species, and we describe its mode of action towards Candida parapsilosis. MaL inhibited the growth of C. albicans and C. parapsilosis. However, MaL was more potent against C. parapsilosis. The candidacidal mode of action of MaL on C. parapsilosis involves enhanced cell permeabilization, alteration of the plasma membrane proton-pumping ATPase function (H+-ATPase), induction of oxidative stress, and DNA damage. MaL also exhibited antibiofilm activity and noncytotoxicity to Vero cells. These results indicate that MaL is a promising candidate for the future development of a new, natural, and safe drug for the treatment of infections caused by C. parapsilosis.
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Antifúngicos/farmacologia , Candida parapsilosis/metabolismo , Estruturas da Membrana Celular/química , Fabaceae/química , Lectinas/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Animais , Antifúngicos/administração & dosagem , Antifúngicos/isolamento & purificação , Apoptose/efeitos dos fármacos , Biofilmes/efeitos dos fármacos , Candida albicans/efeitos dos fármacos , Candida albicans/metabolismo , Candida parapsilosis/citologia , Candida parapsilosis/efeitos dos fármacos , Morte Celular/efeitos dos fármacos , Estruturas da Membrana Celular/metabolismo , Chlorocebus aethiops , Meios de Cultura/análise , Meios de Cultura/química , Dano ao DNA , Lectinas/administração & dosagem , Lectinas/isolamento & purificação , Microscopia Eletrônica de Varredura , Propídio/metabolismo , Sementes/química , Células VeroRESUMO
KEY MESSAGE: Cowpea miRNAs and Argonaute genes showed differential expression patterns in response to CPSMV challenge Several biotic stresses affect cowpea production and yield. CPSMV stands out for causing severe negative impacts on cowpea. Plants have two main induced immune systems. In the basal system (PTI, PAMP-triggered immunity), plants recognize and respond to conserved molecular patterns associated with pathogens (PAMPs). The second type (ETI, Effector-triggered immunity) is induced after plant recognition of specific factors from pathogens. RNA silencing is another important defense mechanism in plants. Our research group has been using biochemical and proteomic approaches to learn which proteins and pathways are involved and could explain why some cowpea genotypes are resistant whereas others are susceptible to CPSMV. This current study was conducted to determine the role of cowpea miRNA in the interaction between a resistant cowpea genotype (BRS-Marataoã) and CPSMV. Previously identified and deposited plant microRNA sequences were used to find out all possible microRNAs in the cowpea genome. This search detected 617 mature microRNAs, which were distributed in 89 microRNA families. Next, 4 out of these 617 miRNAs and their possible target genes that encode the proteins Kat-p80, DEAD-Box, GST, and SPB9, all involved in the defense response of cowpea to CPSMV, had their expression compared between cowpea leaves uninoculated and inoculated with CPSMV. Additionally, the differential expression of genes that encode the Argonaute (AGO) proteins 1, 2, 4, 6, and 10 is reported. In summary, the studied miRNAs and AGO 2 and AGO4 associated genes showed differential expression patterns in response to CPSMV challenge, which indicate their role in cowpea defense.
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Comovirus/fisiologia , Regulação da Expressão Gênica de Plantas , MicroRNAs/genética , Vigna/genética , Vigna/virologia , Sequência de Bases , Genoma de Planta , MicroRNAs/metabolismo , Conformação de Ácido Nucleico , Doenças das Plantas/genética , Doenças das Plantas/virologia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Estabilidade de RNA/genética , Padrões de ReferênciaRESUMO
KEY MESSAGE: SBTX has defensive role against C. kikuchii, and therefore, its constituent genes SBTX17 and SBTX27 are promising candidates to engineer pathogen resistant plants. Soybean (Glycine max [L.] Merr.) is economically the most important legume crop in the world. Its productivity is strongly affected by fungal diseases, which reduce soybean production and seed quality and cause losses of billions of dollars worldwide. SBTX is a protein that apparently takes part in the defensive chemical arsenal of soybean against pathogens. This current study provides data that reinforce this hypothesis. Indeed, SBTX inhibited in vitro the mycelial growth of Cercospora kikuchii, it is constitutively located in the epidermal region of the soybean seed cotyledons, and it is exuded from mature imbibed seeds. Moreover, RT-qPCR analysis of the SBTX associated genes, SBTX17 and SBTX27, which encode for the 17 and 27 kDa polypeptide chains, showed that both genes are expressed in all studied plant tissues during the soybean development, with the highest levels found in the mature seeds and unifoliate leaves. In addition, to assess a local response of the soybean secondary leaves from 35-day-old plants, they were inoculated with C. kikuchii and treated with salicylic acid. It was verified using RT-qPCR that SBTX17 and SBTX27 genes overexpressed in leaves compared to controls. These findings strongly suggest that SBTX has defensive roles against C. kikuchii. Therefore, SBTX17 and SBTX27 genes are promising candidates to engineer pathogen resistant plants.
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Ascomicetos , Resistência à Doença/genética , Glycine max/metabolismo , Glicoproteínas/fisiologia , Doenças das Plantas/microbiologia , Ácido Salicílico/farmacologia , Proteínas de Soja/fisiologia , Ascomicetos/efeitos dos fármacos , Ascomicetos/crescimento & desenvolvimento , Cotilédone/genética , Cotilédone/metabolismo , Expressão Gênica , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Regulação da Expressão Gênica de Plantas/genética , Glicoproteínas/genética , Glicoproteínas/metabolismo , Glicoproteínas/farmacologia , Folhas de Planta/efeitos dos fármacos , Folhas de Planta/genética , Folhas de Planta/metabolismo , Folhas de Planta/microbiologia , Regiões Promotoras Genéticas , Sementes/genética , Sementes/metabolismo , Proteínas de Soja/genética , Proteínas de Soja/metabolismo , Proteínas de Soja/farmacologia , Glycine max/genética , Glycine max/crescimento & desenvolvimento , Glycine max/microbiologia , Regulação para CimaRESUMO
Candida albicans has emerged as a major public health problem in recent decades. The most important contributing factor is the rapid increase in resistance to conventional drugs worldwide. Synthetic antimicrobial peptides (SAMPs) have attracted substantial attention as alternatives and/or adjuvants in therapeutic treatments due to their strong activity at low concentrations without apparent toxicity. Here, two SAMPs, named Mo-CBP3 -PepI (CPAIQRCC) and Mo-CBP3 -PepII (NIQPPCRCC), are described, bioinspired by Mo-CBP3 , which is an antifungal chitin-binding protein from Moringa oleifera seeds. Furthermore, the mechanism of anticandidal activity was evaluated as well as their synergistic effects with nystatin. Both peptides induced the production of reactive oxygen species (ROS), cell wall degradation, and large pores in the C. albicans cell membrane. In addition, the peptides exhibited high potential as adjuvants because of their synergistic effects, by increasing almost 50-fold the anticandidal activity of the conventional antifungal drug nystatin. These peptides have excellent potential as new drugs and/or adjuvants to conventional drugs for treatment of clinical infections caused by C. albicans.
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Antifúngicos/farmacologia , Candida albicans/efeitos dos fármacos , Elétrons , Nistatina/farmacologia , Peptídeos/farmacologia , Antifúngicos/síntese química , Antifúngicos/química , Dicroísmo Circular , Eritrócitos/efeitos dos fármacos , Humanos , Testes de Sensibilidade Microbiana , Microscopia Eletrônica de Varredura , Microscopia de Fluorescência , Nistatina/síntese química , Nistatina/química , Peptídeos/síntese química , Peptídeos/químicaRESUMO
Plant seeds can exudate active molecules with inhibitory effects against several soil pathogens, including nematodes. This study aimed to characterize and evaluate the nematicidal properties against Meloidogyne incognita of exuded proteins from Moringa oleifera seeds. M. oleifera seeds were soaked in distilled water, and exudates were harvested and analyzed for the presence of defense proteins and anthelmintic activity. Enzymatic assays revealed the existence of PR-proteins such as ß-1,3-glucanases (0.18 ± 0.003 nkatal mg-1 of protein), chitinases (0.22 ± 0.004 nkatal mg-1 of protein), proteases (261.30 ± 6.405 AU mg-1 of protein min-1), serine (190.30 ± 5.574 IA mg-1 of protein) and cysteine (231.70 ± 0.923 IA mg-1 of protein), protease inhibitors. The exuded proteins presented ovicidal activity and caused 100% mortality of second-stage juveniles (J2s). Scanning electron microscopy (SEM) revealed deleterious effects on M. incognita eggs, such as invaginations, cracks, scratched surface, and loss of internal content. These findings confirm the presence of anthelmintic proteins in M. oleifera seed exudate, possibly involved in plant defense during seed germination. Besides this, the exuded proteins exhibited strong biotechnological potential for use in the biocontrol of M. incognita infections, which are responsible for millions of dollars in staple crop losses every year.