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1.
Nat Commun ; 12(1): 296, 2021 01 12.
Artigo em Inglês | MEDLINE | ID: mdl-33436600

RESUMO

Nonribosomal peptide synthetases containing starter condensation domains direct the biosynthesis of nonribosomal lipopeptides, which generally exhibit wide bioactivities. The acyl chain has strong impacts on bioactivity and toxicity, but the lack of an in-depth understanding of starter condensation domain-mediated lipoinitiation limits the bioengineering of NRPSs to obtain novel derivatives with desired acyl chains. Here, we show that the acyl chains of the lipopeptides rhizomide, holrhizin, and glidobactin were modified by engineering the starter condensation domain, suggesting a workable approach to change the acyl chain. Based on the structure of the mutated starter condensation domain of rhizomide biosynthetic enzyme RzmA in complex with octanoyl-CoA and related point mutation experiments, we identify a set of residues responsible for the selectivity of substrate acyl chains and extend the acyl chains from acetyl to palmitoyl. Furthermore, we illustrate three possible conformational states of starter condensation domains during the reaction cycle of the lipoinitiation process. Our studies provide further insights into the mechanism of lipoinitiation and the engineering of nonribosomal peptide synthetases.


Assuntos
Lipídeos/química , Biossíntese de Peptídeos Independentes de Ácido Nucleico , Engenharia de Proteínas , Acilação , Sequência de Aminoácidos , Lipopeptídeos/química , Lipopeptídeos/metabolismo , Modelos Moleculares , Mutação Puntual/genética , Domínios Proteicos , Especificidade por Substrato
3.
mBio ; 11(5)2020 10 20.
Artigo em Inglês | MEDLINE | ID: mdl-33082259

RESUMO

The emergence of severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2), the etiological agent of the 2019 coronavirus disease (COVID-19), has erupted into a global pandemic that has led to tens of millions of infections and hundreds of thousands of deaths worldwide. The development of therapeutics to treat infection or as prophylactics to halt viral transmission and spread is urgently needed. SARS-CoV-2 relies on structural rearrangements within a spike (S) glycoprotein to mediate fusion of the viral and host cell membranes. Here, we describe the development of a lipopeptide that is derived from the C-terminal heptad repeat (HRC) domain of SARS-CoV-2 S that potently inhibits infection by SARS-CoV-2. The lipopeptide inhibits cell-cell fusion mediated by SARS-CoV-2 S and blocks infection by live SARS-CoV-2 in Vero E6 cell monolayers more effectively than previously described lipopeptides. The SARS-CoV-2 lipopeptide exhibits broad-spectrum activity by inhibiting cell-cell fusion mediated by SARS-CoV-1 and Middle East respiratory syndrome coronavirus (MERS-CoV) and blocking infection by live MERS-CoV in cell monolayers. We also show that the SARS-CoV-2 HRC-derived lipopeptide potently blocks the spread of SARS-CoV-2 in human airway epithelial (HAE) cultures, an ex vivo model designed to mimic respiratory viral propagation in humans. While viral spread of SARS-CoV-2 infection was widespread in untreated airways, those treated with SARS-CoV-2 HRC lipopeptide showed no detectable evidence of viral spread. These data provide a framework for the development of peptide therapeutics for the treatment of or prophylaxis against SARS-CoV-2 as well as other coronaviruses.IMPORTANCE SARS-CoV-2, the causative agent of COVID-19, continues to spread globally, placing strain on health care systems and resulting in rapidly increasing numbers of cases and mortalities. Despite the growing need for medical intervention, no FDA-approved vaccines are yet available, and treatment has been limited to supportive therapy for the alleviation of symptoms. Entry inhibitors could fill the important role of preventing initial infection and preventing spread. Here, we describe the design, synthesis, and evaluation of a lipopeptide that is derived from the HRC domain of the SARS-CoV-2 S glycoprotein that potently inhibits fusion mediated by SARS-CoV-2 S glycoprotein and blocks infection by live SARS-CoV-2 in both cell monolayers (in vitro) and human airway tissues (ex vivo). Our results highlight the SARS-CoV-2 HRC-derived lipopeptide as a promising therapeutic candidate for SARS-CoV-2 infections.


Assuntos
Antivirais/farmacologia , Betacoronavirus/efeitos dos fármacos , Lipopeptídeos/farmacologia , Glicoproteína da Espícula de Coronavírus/química , Internalização do Vírus/efeitos dos fármacos , Sequência de Aminoácidos , Animais , Antivirais/química , Betacoronavirus/química , Betacoronavirus/fisiologia , Chlorocebus aethiops , Infecções por Coronavirus/prevenção & controle , Infecções por Coronavirus/transmissão , Células HEK293 , Humanos , Lipopeptídeos/química , Fusão de Membrana/efeitos dos fármacos , Coronavírus da Síndrome Respiratória do Oriente Médio/química , Coronavírus da Síndrome Respiratória do Oriente Médio/efeitos dos fármacos , Coronavírus da Síndrome Respiratória do Oriente Médio/fisiologia , Pandemias/prevenção & controle , Pneumonia Viral/prevenção & controle , Pneumonia Viral/transmissão , Domínios Proteicos , Mucosa Respiratória/efeitos dos fármacos , Mucosa Respiratória/virologia , Vírus da SARS/química , Vírus da SARS/efeitos dos fármacos , Vírus da SARS/fisiologia , Células Vero
4.
Proc Natl Acad Sci U S A ; 117(38): 23802-23806, 2020 09 22.
Artigo em Inglês | MEDLINE | ID: mdl-32868430

RESUMO

The bacterial pathogen Pseudomonas tolaasii severely damages white button mushrooms by secretion of the pore-forming toxin tolaasin, the main virulence factor of brown blotch disease. Yet, fungus-associated helper bacteria of the genus Mycetocola (Mycetocola tolaasinivorans and Mycetocola lacteus) may protect their host by an unknown detoxification mechanism. By a combination of metabolic profiling, imaging mass spectrometry, structure elucidation, and bioassays, we found that the helper bacteria inactivate tolaasin by linearizing the lipocyclopeptide. Furthermore, we found that Mycetocola spp. impair the dissemination of the pathogen by cleavage of the lactone ring of pseudodesmin. The role of pseudodesmin as a major swarming factor was corroborated by identification and inactivation of the corresponding biosynthetic gene cluster. Activity-guided fractionation of the Mycetocola proteome, matrix-assisted laser desorption/ionization (MALDI) analyses, and heterologous enzyme production identified the lactonase responsible for toxin cleavage. We revealed an antivirulence strategy in the context of a tripartite interaction that has high ecological and agricultural relevance.


Assuntos
Actinobacteria , Agaricus , Proteínas de Bactérias , Depsipeptídeos , Pseudomonas , Fatores de Virulência , Actinobacteria/química , Actinobacteria/enzimologia , Actinobacteria/fisiologia , Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Depsipeptídeos/química , Depsipeptídeos/metabolismo , Lipopeptídeos/química , Lipopeptídeos/metabolismo , Proteoma , Pseudomonas/química , Pseudomonas/patogenicidade , Fatores de Virulência/química , Fatores de Virulência/metabolismo
5.
Appl Environ Microbiol ; 86(21)2020 10 15.
Artigo em Inglês | MEDLINE | ID: mdl-32859591

RESUMO

In the present study, a deep-sea bacterial strain designated Bacillus sp. strain wsm-1 was screened and found to exhibit strong antifungal activity against many plant-pathogenic fungi, and corresponding antifungal agents were thereby purified and determined by tandem mass spectrometry to be two cyclic lipopeptide homologs. These homologs, which were different from any previously reported lipopeptides, were identified to possess identical amino acid sequences of ß-amino fatty acid-Asn-Ser-Asn-Pro-Tyr-Asn-Gln and deduced as two novel lipopeptides designated C14 iturin W and C15 iturin W. Electron microscopy observation indicated that both iturin W homologs caused obvious morphological changes and serious disruption of plasma membrane toward fungal cells, while C15 iturin W exhibited more serious cell damages than C14 iturin W did, which was well consistent with the results of the antifungal activity assays. To improve the yield and antifungal activity of iturin W, the effects of different carbon and nitrogen sources and amino acids on production of C14 iturin W and C15 iturin W were investigated. The results indicated that supplements of most of the detected carbon and nitrogen sources could increase the yield of C14 iturin W, but inhibit the yield of C15 iturin W, while supplements of tryptone and most of the detected amino acids could increase the yield of both C14 iturin W and C15 iturin W.IMPORTANCE Plant disease caused by pathogenic fungi is one of the most devastating diseases, which affects the food safety of the whole world to a great extent. Biological control of plant diseases by microbial natural products is more desirable than traditional chemical control. In this study, we discovered a novel lipopeptide, iturin W, with promising prospects in biological control of plant diseases. Moreover, the effects of different carbon and nitrogen sources and amino acids on production of C14 iturin W and C15 iturin W would provide a reasonable basis for the optimization of the fermentation process of lipopeptides. Notably, the structure of iturin W was different from that of any previously reported lipopeptide, suggesting that deep-sea microorganisms might produce many novel natural products and have significant potential in the development of biological products in the future.


Assuntos
Antifúngicos/farmacologia , Proteínas de Bactérias/farmacologia , Fungos/efeitos dos fármacos , Lipopeptídeos/farmacologia , Peptídeos Cíclicos/fisiologia , Alternaria/efeitos dos fármacos , Antifúngicos/química , Bacillus , Proteínas de Bactérias/química , Colletotrichum/efeitos dos fármacos , Fungicidas Industriais/química , Fungicidas Industriais/farmacologia , Fusarium/efeitos dos fármacos , Lipopeptídeos/química , Magnaporthe/efeitos dos fármacos , Peptídeos Cíclicos/química , Análise de Sequência de Proteína
6.
Int J Nanomedicine ; 15: 4021-4047, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32606662

RESUMO

Purpose: Periodontitis is a chronic inflammatory disease associated with microbial accumulation. The purpose of this study was to reuse the agricultural waste to produce cellulose nanofibers (CNF) and further modification of the CNF with κ-carrageenan oligosaccharides (CO) for drug delivery. In addition, this study is focused on the antimicrobial activity of surfactin-loaded CO-CNF towards periodontal pathogens. Materials and Methods: A chemo-mechanical method was used to extract the CNF and the modification was done by using CO. The studies were further proceeded by adding different quantities of surfactin [50 mg (50 SNPs), 100 mg (100 SNPs), 200 mg (200 SNPs)] into the carrier (CO-CNF). The obtained materials were characterized, and the antimicrobial activity of surfactin-loaded CO-CNF was evaluated. Results: The obtained average size of CNF and CO-CNF after ultrasonication was 263 nm and 330 nm, respectively. Microscopic studies suggested that the CNF has a short diameter with long length and CO became cross-linked to form as beads within the CNF network. The addition of CO improved the degradation temperature, crystallinity, and swelling property of CNF. The material has a controlled drug release, and the entrapment efficiency and loading capacity of the drug were 53.15 ± 2.36% and 36.72 ± 1.24%, respectively. It has antioxidant activity and inhibited the growth of periodontal pathogens such as Streptococcus mutans and Porphyromonas gingivalis by preventing the biofilm formation, reducing the metabolic activity, and promoting the oxidative stress. Conclusion: The study showed the successful extraction of CNF and modification with CO improved the physical parameters of the CNF. In addition, surfactin-loaded CO-CNF has potential antimicrobial activity against periodontal pathogens. The obtained biomaterial is economically valuable and has great potential for biomedical applications.


Assuntos
Carragenina/química , Celulose/química , Lipopeptídeos/química , Nanofibras/química , Peptídeos Cíclicos/química , Periodonto/microbiologia , Animais , Bactérias/metabolismo , Compostos de Bifenilo/química , Sobrevivência Celular , Difusão Dinâmica da Luz , Depuradores de Radicais Livres/química , Malondialdeído/metabolismo , Camundongos , Testes de Sensibilidade Microbiana , Nanofibras/ultraestrutura , Oligossacarídeos/química , Picratos/química , Células RAW 264.7 , Soja/química , Espectroscopia de Infravermelho com Transformada de Fourier
7.
J Virol ; 94(14)2020 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-32376627

RESUMO

The 2019 coronavirus disease (COVID-19), caused by the emerging severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has posed serious threats to global public health and economic and social stabilities, calling for the prompt development of therapeutics and prophylactics. In this study, we first verified that SARS-CoV-2 uses human angiotensin-converting enzyme 2 (ACE2) as a cell receptor and that its spike (S) protein mediates high membrane fusion activity. The heptad repeat 1 (HR1) sequence in the S2 fusion protein of SARS-CoV-2 possesses markedly increased α-helicity and thermostability, as well as a higher binding affinity with its corresponding heptad repeat 2 (HR2) site, than the HR1 sequence in S2 of severe acute respiratory syndrome coronavirus (SARS-CoV). Then, we designed an HR2 sequence-based lipopeptide fusion inhibitor, termed IPB02, which showed highly potent activities in inhibiting SARS-CoV-2 S protein-mediated cell-cell fusion and pseudovirus transduction. IPB02 also inhibited the SARS-CoV pseudovirus efficiently. Moreover, the structure-activity relationship (SAR) of IPB02 was characterized with a panel of truncated lipopeptides, revealing the amino acid motifs critical for its binding and antiviral capacities. Therefore, the results presented here provide important information for understanding the entry pathway of SARS-CoV-2 and the design of antivirals that target the membrane fusion step.IMPORTANCE The COVID-19 pandemic, caused by SARS-CoV-2, presents a serious global public health emergency in urgent need of prophylactic and therapeutic interventions. The S protein of coronaviruses mediates viral receptor binding and membrane fusion, thus being considered a critical target for antivirals. Herein, we report that the SARS-CoV-2 S protein has evolved a high level of activity to mediate cell-cell fusion, significantly differing from the S protein of SARS-CoV that emerged previously. The HR1 sequence in the fusion protein of SARS-CoV-2 adopts a much higher helical stability than the HR1 sequence in the fusion protein of SARS-CoV and can interact with the HR2 site to form a six-helical bundle structure more efficiently, underlying the mechanism of the enhanced fusion capacity. Also, importantly, the design of membrane fusion inhibitors with high potencies against both SARS-CoV-2 and SARS-CoV has provided potential arsenals to combat the pandemic and tools to exploit the fusion mechanism.


Assuntos
Antivirais/farmacologia , Betacoronavirus/efeitos dos fármacos , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/virologia , Lipopeptídeos/farmacologia , Fusão de Membrana/efeitos dos fármacos , Pneumonia Viral/tratamento farmacológico , Pneumonia Viral/virologia , Sequência de Aminoácidos , Betacoronavirus/fisiologia , Desenho de Fármacos , Células HEK293 , Humanos , Lipopeptídeos/química , Glicoproteínas de Membrana/metabolismo , Pandemias , Peptidil Dipeptidase A/metabolismo , Ligação Proteica , Glicoproteína da Espícula de Coronavírus/antagonistas & inibidores , Glicoproteína da Espícula de Coronavírus/metabolismo , Proteínas do Envelope Viral/metabolismo
8.
J Med Chem ; 63(11): 6090-6095, 2020 06 11.
Artigo em Inglês | MEDLINE | ID: mdl-32378891

RESUMO

A novel lipopeptide antibiotic, stalobacin I (1), was discovered from a culture broth of an unidentified Gram-negative bacterium. Stalobacin I (1) had a unique chemical architecture composed of an upper and a lower half peptide sequence, which were linked via a hemiaminal methylene moiety. The sequence of 1 contained an unusual amino acid, carnosadine, 3,4-dihydroxyariginine, 3-hydroxyisoleucine, and 3-hydroxyaspartic acid, and a novel cyclopropyl fatty acid. The antibacterial activity of 1 against a broad range of drug-resistant Gram-positive bacteria was much stronger than those of "last resort" antibiotics such as vancomycin, linezolid, and telavancin (MIC 0.004-0.016 µg/mL). Furthermore, compound 1 induced a characteristic morphological change in Gram-positive and Gram-negative strains by inflating the bacterial cell body. The absolute configuration of a cyclopropyl amino acid, carnosadine, was determined by the synthetic study of its stereoisomers, which was an essential component for the strong activity of 1.


Assuntos
Antibacterianos/farmacologia , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Lipopeptídeos/química , Aminoglicosídeos/farmacologia , Antibacterianos/química , Avaliação Pré-Clínica de Medicamentos , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Positivas/efeitos dos fármacos , Lipoglicopeptídeos/farmacologia , Lipopeptídeos/farmacologia , Testes de Sensibilidade Microbiana
9.
Sci Rep ; 10(1): 7269, 2020 04 29.
Artigo em Inglês | MEDLINE | ID: mdl-32350300

RESUMO

Lipidised analgesic peptide prodrugs self-assemble into peptide nanofibers; with the nanofiber morphology protecting the peptide from plasma degradation and improving therapeutic efficacy. Extending this learning, we hypothesised that a self-assembling lipidized peptide arginine vasopressin (AVP) receptor agonist, that had not been designed as a prodrug, could prove pharmacologically active and control urine production. The only approved AVP receptor agonist, desmopressin is indicated for the treatment of central diabetes insipidus (DI), bedwetting, haemophilia A and von Willebrand disease. Desmopressin is well tolerated by most patients, however adverse effects, such as hyponatraemia and water intoxication necessitate a strict fluid intake, thus motivating the search for alternative DI treatments. Selective V2 receptor agonism is required for anti-DI activity and we hypothesised that our new lipidized peptide (METx) would lead to selective AVP receptor agonism. METx was synthesised and characterised and then tested for activity against the V2, V1a and OT uterine receptors and not tested against the V1b receptor as METx was not expected to cross the blood brain barrier. METx was also tested in vivo in a healthy rat model. METx forms nanofibers and is a partial V2 receptor agonist (determined by measuring MDCK cell line cAMP accumulation), producing 57% of AVP's maximal activity (EC50 = 2.7 nM) and is not a V1a agonist up to a concentration of 1 µM (determined by measuring A7r5 cell line D-myo-inositol-1-phosphate accumulation). METx is a weak OT receptor antagonist, reducing the frequency of OT induced contractions (EC50 = 350 nM) and increasing the OT EC50 from 0.081 nM to 21 nM at a concentration of 600 nM. METx (41 nM) had no effect on spontaneous uterine contractions and METx (100 nM) had no effect on OT induced uterine contractions. Simulated binding studies show that binding avidity to the receptors follows the trend: V2 > OT > V1a. On intravenous injection, a nanoparticle formulation of METx reduced urine production in a healthy rat model in a dose responsive manner, with 40 mg kg-1 METx resulting in no urine production over 4 hours. The lipidized self-assembling peptide - METx - is a selective competitive V2 receptor agonist and an anti-diuretic.


Assuntos
Antidiuréticos , Arginina Vasopressina , Lipopeptídeos , Receptores de Vasopressinas/agonistas , Urina , Animais , Antidiuréticos/síntese química , Antidiuréticos/química , Antidiuréticos/farmacologia , Arginina Vasopressina/síntese química , Arginina Vasopressina/química , Arginina Vasopressina/farmacologia , Cães , Feminino , Lipopeptídeos/síntese química , Lipopeptídeos/química , Lipopeptídeos/farmacologia , Células Madin Darby de Rim Canino , Masculino , Ratos , Ratos Sprague-Dawley , Ratos Wistar , Receptores de Vasopressinas/metabolismo
10.
J Virol ; 94(15)2020 07 16.
Artigo em Inglês | MEDLINE | ID: mdl-32404526

RESUMO

We recently reported a group of lipopeptide-based membrane fusion inhibitors with potent antiviral activities against human immunodeficiency virus type 1 (HIV-1), HIV-2, and simian immunodeficiency virus (SIV). In this study, the in vivo therapeutic efficacy of such a lipopeptide, LP-52, was evaluated in rhesus macaques chronically infected with pathogenic SIVmac239. In a pilot study with one monkey, monotherapy with low-dose LP-52 rapidly reduced the plasma viral loads to below the limit of detection and maintained viral suppression during three rounds of structurally interrupted treatment. The therapeutic efficacy of LP-52 was further verified in four infected monkeys; however, three out of the monkeys had viral rebounds under the LP-52 therapy. We next focused on characterizing SIV mutants responsible for the in vivo resistance. Sequence analyses revealed that a V562A or V562M mutation in the N-terminal heptad repeat (NHR) and a E657G mutation in the C-terminal heptad repeat (CHR) of SIV gp41 conferred high resistance to LP-52 and cross-resistance to the peptide drug T20 and two newly designed lipopeptides (LP-80 and LP-83). Moreover, we showed that the resistance mutations greatly reduced the stability of diverse fusion inhibitors with the NHR site, and V562A or V562M in combination with E657G could significantly impair the functionality of viral envelopes (Envs) to mediate SIVmac239 infection and decrease the thermostability of viral six-helical bundle (6-HB) core structure. In conclusion, the present data have not only facilitated the development of novel anti-HIV drugs that target the membrane fusion step, but also help our understanding of the mechanism of viral evolution to develop drug resistance.IMPORTANCE The anti-HIV peptide drug T20 (enfuvirtide) is the only membrane fusion inhibitor available for treatment of viral infection; however, it exhibits relatively weak antiviral activity, short half-life, and a low genetic barrier to inducing drug resistance. Design of lipopeptide-based fusion inhibitors with extremely potent and broad antiviral activities against divergent HIV-1, HIV-2, and SIV isolates have provided drug candidates for clinical development. Here, we have verified a high therapeutic efficacy for the lipopeptide LP-52 in SIVmac239-infected rhesus monkeys. The resistance mutations selected in vivo have also been characterized, providing insights into the mechanism of action of newly designed fusion inhibitors with a membrane-anchoring property. For the first time, the data show that HIV-1 and SIV can share a similar genetic pathway to develop resistance, and that a lipopeptide fusion inhibitor could have a same resistance profile as its template peptide.


Assuntos
Lipopeptídeos/farmacologia , Lipoproteínas/farmacologia , Síndrome de Imunodeficiência Adquirida dos Símios , Vírus da Imunodeficiência Símia/metabolismo , Proteínas Virais de Fusão/metabolismo , Internalização do Vírus/efeitos dos fármacos , Substituição de Aminoácidos , Animais , Lipopeptídeos/química , Lipoproteínas/química , Macaca mulatta , Mutação de Sentido Incorreto , Síndrome de Imunodeficiência Adquirida dos Símios/tratamento farmacológico , Síndrome de Imunodeficiência Adquirida dos Símios/genética , Síndrome de Imunodeficiência Adquirida dos Símios/metabolismo , Síndrome de Imunodeficiência Adquirida dos Símios/patologia , Vírus da Imunodeficiência Símia/genética , Proteínas Virais de Fusão/genética
11.
J Med Chem ; 63(10): 5387-5397, 2020 05 28.
Artigo em Inglês | MEDLINE | ID: mdl-32347723

RESUMO

Group A Streptococcus (GAS) infection causes a range of life-threatening diseases, including rheumatic heart disease. Cyclic peptides offer an attractive solution for presentation of short peptide antigens due to their stability and structurally constrained conformation. We investigated a cyclic carrier decapeptide incorporating a B cell GAS peptide epitope, a universal T helper epitope, and a synthetic toll-like receptor 2-targeting moiety as a possible self-adjuvanting GAS vaccine. A structure-activity relationship of the cyclic lipopeptide vaccine showed successful induction of J8-specific systemic immunoglobulin G (IgG) antibodies when administered subcutaneously without an additional adjuvant. Interestingly, the physical mixture control induced the highest titers of all vaccine compounds, with antibodies from mice immunized with this physical mixture control shown to effectively opsonize multiple strains of clinically isolated GAS bacteria. This study showed the capability for a self-adjuvanting cyclic delivery system to act as a vehicle for the delivery of GAS peptide antigens to treat GAS infections.


Assuntos
Antibacterianos/metabolismo , Antígenos de Bactérias/metabolismo , Vacinas Bacterianas/metabolismo , Lipopeptídeos/metabolismo , Infecções Estreptocócicas/metabolismo , Streptococcus pyogenes/metabolismo , Animais , Antibacterianos/administração & dosagem , Antibacterianos/química , Vacinas Bacterianas/administração & dosagem , Vacinas Bacterianas/química , Quimioterapia Adjuvante/métodos , Feminino , Humanos , Lipopeptídeos/administração & dosagem , Lipopeptídeos/química , Camundongos , Camundongos Endogâmicos C57BL , Infecções Estreptocócicas/tratamento farmacológico , Streptococcus pyogenes/efeitos dos fármacos , Relação Estrutura-Atividade
12.
Toxicol Lett ; 328: 1-6, 2020 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-32315709

RESUMO

The genotoxicity of cationic lipopeptide nanoparticles (cLPNPs) was evaluated in vivo and in vitro comet assay and the in vivo chromosome aberrations test. In vitro comet assay, human blood cells were exposed to cLPNPs at the concentration of 2.5, 5, 10, 20, 40 and 100 µg/mL. Significant DNA damage was observed after 1 h exposure, but no effects were detected after 3 h. In vivo, cLPNPs were administered in single or five daily injection doses at 8, 20 and 40 mg/kg of body weight by subcutaneous injection to male mice. The cLPNPs caused DNA damage in the liver, lung and kidney, but not in the spleen. The kidney was more prone to genotoxic effects that persisted from 24 h to 14d after a single injection of cLPNPs. No statistically significant increase in the percentage of cells with chromosomal aberrations above the vehicle control was observed in mice bone marrow after a single or repeated injection of cLPNPs. In summary, cLPNPs shown to be genotoxic both in vivo and in vitro. The results suggest the importance of the use of highly sensitive methods, such as the comet assay, in order to determine the full genotoxic potential of nanoparticles.


Assuntos
Aberrações Cromossômicas/induzido quimicamente , Dano ao DNA , Lipopeptídeos/toxicidade , Nanopartículas/toxicidade , Animais , Células da Medula Óssea/efeitos dos fármacos , Células da Medula Óssea/patologia , Sobrevivência Celular/efeitos dos fármacos , Ensaio Cometa , Relação Dose-Resposta a Droga , Humanos , Injeções Subcutâneas , Rim/efeitos dos fármacos , Rim/patologia , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/patologia , Lipopeptídeos/química , Fígado/efeitos dos fármacos , Fígado/patologia , Pulmão/efeitos dos fármacos , Pulmão/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , Nanopartículas/química
13.
Drug Des Devel Ther ; 14: 541-550, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32103896

RESUMO

Background: Microemulsions drug delivery systems (MDDS) have been known to increase the bioavailability of hydrophobic drugs. The main challenge of the MDDS is the development of an effective and safe system for drug carriage and delivery. Biosurfactants are preferred surface-active molecules because of their lower toxicity and safe characteristics when compared to synthetic surfactants. Glycolipid and lipopeptide are the most common biosurfactants that were tested for MDDS. The main goal of the present systematic review was to estimate the available evidence on the role of biosurfactant in the development of MDDS. Search Strategy: Literature searches involved the main scientific databases and were focused on the period from 2005 until 2017. The Search filter composed of two items: "Biosurfactant" and/or "Microemulsion." Inclusion Criteria: Twenty-four studies evaluating the use of biosurfactant in MDDS were eligible for inclusion. Among these 14 were related to the use of glycolipid biosurfactants in the MDDS formulations, while four reported using lipopeptide biosurfactants and six other related review articles. Results: According to the output study parameters, biosurfactants acted as active stabilizers, hydrophilic or hydrophobic linkers and safety carriers in MDDS, and among them glycolipid biosurfactants had the most application in MDDS formulations. Conclusion: Synthetic surfactants could be replaced by biosurfactants as an effective bio-source for MDDS due to their excellent self-assembling and emulsifying activity properties.


Assuntos
Sistemas de Liberação de Medicamentos , Preparações Farmacêuticas/administração & dosagem , Tensoativos/química , Portadores de Fármacos/química , Emulsões , Glicolipídeos/química , Humanos , Interações Hidrofóbicas e Hidrofílicas , Lipopeptídeos/química , Preparações Farmacêuticas/química
14.
Planta ; 251(3): 70, 2020 Feb 21.
Artigo em Inglês | MEDLINE | ID: mdl-32086615

RESUMO

MAIN CONCLUSION: Lipopeptides could help to overcome a large concern in agriculture: resistance against chemical pesticides. These molecules have activity against various phytopathogens and a potential to be transformed by genetic engineering. The exponential rise of pest resistances to different chemical pesticides and the global appeal of consumers for a sustainable agriculture and healthy nutrition have led to the search of new solutions for pest control. Furthermore, new laws require a different stance of producers. Based on that, bacteria of the genus Bacillus present a great agricultural potential, producing lipopeptides (LPs) that have high activity against insects, mites, nematodes, and/or phytopathogens that are harmful to plant cultures. Biopesticide activity can be found mainly in three families of Bacillus lipopeptides: surfactin, iturin, and fengycin. These molecules have an amphiphilic nature, interfering with biological membrane structures. Their antimicrobial properties include activity against bacteria, fungi, oomycetes, and viruses. Recent studies also highlight the ability of these compounds to stimulate defense mechanisms of plants and biofilm formation, which is a key factor for the successful colonization of biocontrol organisms. The use of molecular biology has also recently been researched for continuous advances and discoveries of new LPs, avoiding possible future problems of resistance against these molecules. As a consequence of the properties and possibilities of LPs, numerous studies and developments as well as the attention of large companies in the field is expected in the near future.


Assuntos
Agricultura , Bacillus/metabolismo , Lipopeptídeos/farmacologia , Controle Biológico de Vetores/métodos , Animais , Anti-Infecciosos/farmacologia , Antifúngicos/farmacologia , Bactérias/efeitos dos fármacos , Resistência a Medicamentos , Fungos/efeitos dos fármacos , Insetos/efeitos dos fármacos , Lipopeptídeos/química , Lipopeptídeos/metabolismo , Ácaros/efeitos dos fármacos , Nematoides/efeitos dos fármacos , Peptídeos Cíclicos/farmacologia , Praguicidas/farmacologia , Doenças das Plantas/prevenção & controle , Plantas/microbiologia , Vírus/efeitos dos fármacos
15.
Molecules ; 25(4)2020 Feb 13.
Artigo em Inglês | MEDLINE | ID: mdl-32069902

RESUMO

Antimicrobial resistance to conventional antibiotics and the limited alternatives to combat plant-threatening pathogens are worldwide problems. Antibiotic lipopeptides exert remarkable membrane activity, which usually is not prone to fast resistance formation, and often show organism-type selectivity. Additional modes of action commonly complement the bioactivity profiles of such compounds. The present work describes a multicomponent-based methodology for the synthesis of cyclic polycationic lipopeptides with stabilized helical structures. The protocol comprises an on solid support Ugi-4-component macrocyclization in the presence of a lipidic isocyanide. Circular dichroism was employed to study the influence of both macrocyclization and lipidation on the amphiphilic helical structure in water and micellar media. First bioactivity studies against model phytopathogens demonstrated a positive effect of the lipidation on the antimicrobial activity.


Assuntos
Antifúngicos/química , Lactamas/química , Lipopeptídeos/química , Peptídeos Cíclicos/química , Antibacterianos/síntese química , Antibacterianos/química , Antibacterianos/farmacologia , Antifúngicos/síntese química , Antifúngicos/farmacologia , Botrytis/efeitos dos fármacos , Lipopeptídeos/síntese química , Lipopeptídeos/farmacologia , Peptídeos Cíclicos/síntese química , Peptídeos Cíclicos/farmacologia , Phytophthora infestans/efeitos dos fármacos
16.
Sci Rep ; 10(1): 2839, 2020 02 18.
Artigo em Inglês | MEDLINE | ID: mdl-32071421

RESUMO

The novel anti-fungal cyclic lipopeptide 'Kannurin' and its three structural variants produced by Bacillus cereus AK1 were previously reported from our laboratory. The present study reports unexplored structural variants of Kannurin those have functional benefits. Due to the difference in ß-hydroxy fatty acid tail length, they are designated here as Kannurin A (m/z 994.67 ± 0.015), B (m/z 1008.68 ± 0.017), C (m/z 1022.69 ± 0.021), D (m/z 1036.70 ± 0.01), CL (m/z 1040.71 ± 0.02) and DL (m/z 1054.72 ± 0.01). The isoform A (m/z 994.67 ± 0.015) is the shortest cyclic form of Kannurin identified so far. In addition, CL (m/z 1040.71 ± 0.02) and DL (m/z 1054.72 ± 0.01) are the rare natural linear forms. The results of the antimicrobial assays deduced that the difference in lipid tail length of the isoforms contributes tremendous differences in their antimicrobial properties. The isoforms with short lipid tails (A and B) are more selective and potent towards bacteria, whereas the isoforms with long lipid tails (C and D) are more potent against fungi. The molecular dynamics studies and electron microscopic observations supported with circular dichroic spectroscopy analysis showed the structural confirmation and formation of aggregates of Kannurin in solution. The molecular dynamics simulation studies revealed that a single molecule of Kannurin makes enormous intra-molecular interactions and structural re-arrangements to attain stable lowest energy state in solution. When they reach a particular concentration (CMC) especially in aqueous environment, tends to form structural aggregates called 'micelles'. With the structural information and activity relationship described in this study, it is trying to point out the sensitive structural entities that can be modified to improve the efficacy and target specificities of lipopeptide class of antibiotics.


Assuntos
Ácidos Graxos/química , Lipídeos/química , Lipopeptídeos/química , Anti-Infecciosos/química , Anti-Infecciosos/farmacologia , Bacillus cereus/química , Bacillus cereus/genética , Ácidos Graxos/genética , Fungos/efeitos dos fármacos , Fungos/patogenicidade , Lipídeos/genética , Lipopeptídeos/genética , Micelas , Isoformas de Proteínas/química , Isoformas de Proteínas/genética
17.
Org Biomol Chem ; 18(9): 1710-1727, 2020 03 04.
Artigo em Inglês | MEDLINE | ID: mdl-32052002

RESUMO

Nonribosomal lipopeptides (NRLPs) are complex natural products of bacterial origin that not only fulfill important ecological functions but also serve as lead structures for the development of new pharmaceuticals. In order to carry out detailed structure-activity relationship studies and to decipher the biological activities of NRLPs, the primary structure, including stereochemical assignment, of every new member of this natural product family has to be established first. In this review, we want to focus on analytical techniques and tools that can be employed to elucidate the structure of bacterial NRLPs.


Assuntos
Proteínas de Bactérias/química , Lipopeptídeos/química , Estrutura Molecular , Biossíntese de Peptídeos Independentes de Ácido Nucleico , Relação Estrutura-Atividade
18.
Molecules ; 25(2)2020 Jan 08.
Artigo em Inglês | MEDLINE | ID: mdl-31936341

RESUMO

Ultrashort cationic lipopeptides (USCLs) are promising antimicrobial agents that hypothetically may be alternatively used to combat pathogens such as bacteria and fungi. In general, USCLs consist of fatty acid chains and a few basic amino acid residues. The main shortcoming of USCLs is their relatively high cytotoxicity and hemolytic activity. This study focuses on the impact of the hydrophobic fatty acid chain, on both antimicrobial and hemolytic activities. To learn more about this region, a series of USCLs with different straight-chain fatty acids (C8, C10, C12, C14) attached to the tripeptide with two arginine residues were synthesized. The amino acid at the N-terminal position was exchanged for proteinogenic and non-proteinogenic amino acid residues (24 in total). Moreover, the branched fatty acid residues were conjugated to N-terminus of a dipeptide with two arginine residues. All USCLs had C-terminal amides. USCLs were tested against reference bacterial strains (including ESKAPE group) and Candida albicans. The hemolytic potential was tested on human erythrocytes. Hydrophobicity of the compounds was evaluated by RP-HPLC. Shortening of the fatty acid chain and simultaneous addition of amino acid residue at N-terminus were expected to result in more selective and active compounds than those of the reference lipopeptides with similar lipophilicity. Hypothetically, this approach would also be beneficial to other antimicrobial peptides where N-lipidation strategy was used to improve their biological characteristics.


Assuntos
Aminoácidos/farmacologia , Antibacterianos/farmacologia , Ácidos Graxos/farmacologia , Hemólise/efeitos dos fármacos , Lipopeptídeos/farmacologia , Cátions , Humanos , Interações Hidrofóbicas e Hidrofílicas , Lipopeptídeos/química , Testes de Sensibilidade Microbiana , Relação Estrutura-Atividade
19.
Sci Total Environ ; 714: 136400, 2020 Apr 20.
Artigo em Inglês | MEDLINE | ID: mdl-31982734

RESUMO

Apparent solubility and bioavailability of hydrophobic compounds are the major problems in the bioremediation process, which could be overcome by the bacteria capable of biosurfactant production and concurrent hydrocarbon degradation. In this work, we constructed an artificial bacterial consortium containing Lysinibacillus, Paenibacillus, Gordonia and Cupriavidus spp. from glyceryl tributyrate enriched bacteria collected from the non-contaminated site. The consortium was capable of using common raw materials (olive oil, paraffin oil, and glycerol) and polyaromatic hydrocarbons pollutants (naphthalene and anthracene) as the sole carbon source with simultaneous biosurfactant production. Two new lipopeptide isoforms, containing heptapeptide and lipid moieties, were structurally elucidated by LC-MS/MS, FTIR, NMR and molecular networking analysis. Our findings indicate that hydrocarbons degradation and biosurfactant production is an intrinsic property of non-contaminated soil community. Interestingly, we observed the hyper chemotactic activity of Lysinibacillus strains towards glyceryl tributyrate, which has not been reported before. The study may deepen our understanding of microbial strains and consortium with the potential to be used for bioremediation of hydrocarbons contaminated environments.


Assuntos
Lipopeptídeos/química , Biodegradação Ambiental , Cromatografia Líquida , Hidrocarbonetos , Petróleo , Microbiologia do Solo , Poluentes do Solo , Tensoativos , Espectrometria de Massas em Tandem
20.
Curr Microbiol ; 77(3): 443-451, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31894376

RESUMO

Fengycin antibiotic displays a strong antifungal activity and inhibits the growth of a wide range of plant pathogens especially filamentous fungi. The main objective of the present study is to characterize fengycin variants produced by B. amyloliquefaciens strain (ET). LC-MS analysis of fengycin extracts has shown several molecular ion peaks corresponding to conventional fengycin homologues (MH + : m/z 1463.9; 1491.9; 1506) and some new ones (MH + : m/z 1433; 1447; 1461; and 1477). Further characterization of these precursor ions was carried out by LC-MS.MS analysis. Reporter fragment ions were observed (named A and B), they correspond to the cleavage of Orn2-Tyr3 (A), Glu1-Orn2 (B), and used for identifying fengycin variants. The reporter fragment couple ions [A/B] at [m/z 966.5/1080.5] and [m/z 994.4 /1108.5] represent fengycin A and B, respectively. The diagnostic ions at ([m/z 980/1094]) may correspond to fengycin C3, D, S or B2. Interestingly, unknown diagnostic product ions at [m/z 951/1065] and [m/z 979/1093] were detected for the first time in this study which prove that they correspond to new fengycin variants, named fengycin X and fengycin Y, respectively. The fengycin X results from a substitution of the glutamine amino acid (Q), at position 8 of the fengycin A peptide part, by an isoleucine (I) or a leucine (L) residue. This mutation should be the same in fengycin Y but compared to fengycin B.


Assuntos
Bacillus amyloliquefaciens/química , Bacillus amyloliquefaciens/genética , Lagos/microbiologia , Lipopeptídeos/química , Argélia , Substituição de Aminoácidos , Glutamina/genética , Isoleucina/genética , Leucina/genética , Mutação , Águas Salinas , Espectrometria de Massas em Tandem
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