RESUMO
Staphylococcus aureus has the ability to invade cortical bone osteocyte lacuno-canalicular networks (OLCNs) and cause osteomyelitis. It was recently established that the cell wall transpeptidase, penicillin-binding protein 4 (PBP4), is crucial for this function, with pbp4 deletion strains unable to invade OLCNs and cause bone pathogenesis in a murine model of S. aureus osteomyelitis. Moreover, PBP4 has recently been found to modulate S. aureus resistance to ß-lactam antibiotics. As such, small molecule inhibitors of S. aureus PBP4 may represent dual functional antimicrobial agents that limit osteomyelitis and/or reverse antibiotic resistance. A high throughput screen recently revealed that the phenyl-urea 1 targets PBP4. Herein, we describe a structure-activity relationship (SAR) study on 1. Leveraging in silico docking and modeling, a set of analogs was synthesized and assessed for PBP4 inhibitory activities. Results revealed a preliminary SAR and identified lead compounds with enhanced binding to PBP4, more potent antibiotic resistance reversal, and diminished PBP4 cell wall transpeptidase activity in comparison to 1.
Assuntos
Antibacterianos , Simulação de Acoplamento Molecular , Proteínas de Ligação às Penicilinas , Staphylococcus aureus , Proteínas de Ligação às Penicilinas/metabolismo , Proteínas de Ligação às Penicilinas/antagonistas & inibidores , Antibacterianos/farmacologia , Antibacterianos/química , Relação Estrutura-Atividade , Staphylococcus aureus/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , Ureia/química , Ureia/farmacologia , Ureia/análogos & derivados , Animais , Camundongos , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/antagonistas & inibidoresRESUMO
Antimicrobial resistance has recently been considered an emerging catastrophe globally. The public health and environmental threats were aggravated by the injudicious use of antibiotics in animal farming, aquaculture, and croup fields, etc. Consequently, failure of antibiotic therapies is common because of the emergence of multidrug-resistant (MDR) bacteria in the environment. Thus, the reduction in antibiotic spillage in the environment could be an important step for overcoming this situation. Bear in mind, this research was focused on the green synthesis of chitosan nanoparticles (ChiNPs) using Citrus lemon (Assam lemon) extract as a cross-linker and application in controlling MDR bacteria to reduce the antibiotic spillage in that sector. For evaluating antibacterial activity, Staphylococcus aureus and Escherichia coli were isolated from environmental specimens, and their multidrug-resistant pattern were identified both phenotypically by disk diffusion and genotypically by detecting methicillin- (mecA), penicillin- (blaZ), and streptomycin (aadA1)-resistance encoding genes. The inhibitory zone's diameter was employed as a parameter for determining the antibacterial effect against MDR bacteria revealing 30 ± 0.4 mm, 34 ± 0.2 mm, and 36 ± 0.8 mm zones of inhibition against methicillin- (mecA) and penicillin (blaZ)-resistant S. aureus, and streptomycin (aadA1)-resistant E. coli, respectively. The minimum inhibitory concentration at 0.31 mg/mL and minimum bactericidal concentration at 0.62 mg/mL of yielded ChiNPs were used as the broad-spectrum application against MDR bacteria. Finally, the biocompatibility of ChiNPs was confirmed by showing a negligible decrease in BHK-21 cell viability at doses less than 2 MIC, suggesting their potential for future application in antibiotic-free farming practices.
Assuntos
Antibacterianos , Quitosana , Farmacorresistência Bacteriana Múltipla , Escherichia coli , Nanopartículas , Staphylococcus aureus , Antibacterianos/farmacologia , Antibacterianos/química , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/genética , Quitosana/farmacologia , Quitosana/química , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Química Verde , Testes de Sensibilidade Microbiana , Nanopartículas/química , Proteínas de Ligação às Penicilinas/genética , Proteínas de Ligação às Penicilinas/metabolismo , Proteínas de Ligação às Penicilinas/antagonistas & inibidores , Staphylococcus aureus/efeitos dos fármacosRESUMO
The penicillin binding protein 2a (PBP2a) is a key enzyme associated with bacterial cell wall synthesis and bacterial infection. Therefore, targeting PBPa2 offers a promising approach for the therapeutics of bacterial resistance and infection. This study presents a comprehensive analysis of alpha-mangostin as a potential inhibitor of PBPa2. Molecular docking simulations revealed a strong binding affinity between alpha-mangostin and PBP2a, with an affinity score of -6.01 kcal/mol. Notably, alpha-mangostin formed a preferential hydrogen bond with THR216 of PBP2a, alongside several other polar and hydrophobic interactions. ADME and Toxicity predictions indicated that alpha-mangostin possesses favourable pharmacokinetic properties, suggesting its potential as a therapeutic agent. PASS analysis further highlighted its broad range of favourable biological properties. SwissTargetPrediction analysis reinforced these findings, indicating alpha-mangostin's association with various biological processes. Cell toxicity assays demonstrated that alpha-mangostin had no significant impact on the viability of HEK-293 cells, suggesting its potential safety for further development. The IC50 value for alpha-mangostin was found to be 33.43µM. Fluorescence-based binding assays showed that alpha-mangostin effectively inhibited PBP2a activity in a concentration-dependent manner, supporting its role as an inhibitor. In conclusion, the results suggest alpha-mangostin as a promising candidate for inhibiting PBP2a. Further, extensive studies are warranted to explore its clinical applications.
Assuntos
Antibacterianos , Staphylococcus aureus Resistente à Meticilina , Simulação de Acoplamento Molecular , Proteínas de Ligação às Penicilinas , Xantonas , Proteínas de Ligação às Penicilinas/antagonistas & inibidores , Proteínas de Ligação às Penicilinas/metabolismo , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Humanos , Xantonas/química , Xantonas/farmacologia , Células HEK293 , Antibacterianos/farmacologia , Antibacterianos/química , Testes de Sensibilidade Microbiana , Infecções Estafilocócicas/tratamento farmacológico , Infecções Estafilocócicas/microbiologia , Bibliotecas de Moléculas Pequenas/farmacologia , Bibliotecas de Moléculas Pequenas/química , Proteínas de Bactérias/antagonistas & inibidores , Proteínas de Bactérias/metabolismo , Ligação ProteicaRESUMO
ß-Lactam antibiotics disrupt the assembly of peptidoglycan (PG) within the bacterial cell wall by inhibiting the enzymatic activity of penicillin-binding proteins (PBPs). It was recently shown that ß-lactam treatment initializes a futile cycle of PG synthesis and degradation, highlighting major gaps in our understanding of the lethal effects of PBP inhibition by ß-lactam antibiotics. Here, we assess the downstream metabolic consequences of treatment of Escherichia coli with the ß-lactam mecillinam and show that lethality from PBP2 inhibition is a specific consequence of toxic metabolic shifts induced by energy demand from multiple catabolic and anabolic processes, including accelerated protein synthesis downstream of PG futile cycling. Resource allocation into these processes is coincident with alterations in ATP synthesis and utilization, as well as a broadly dysregulated cellular redox environment. These results indicate that the disruption of normal anabolic-catabolic homeostasis by PBP inhibition is an essential factor for ß-lactam antibiotic lethality.
Assuntos
Andinocilina/farmacologia , Antibacterianos/farmacologia , Proteínas de Escherichia coli/antagonistas & inibidores , Escherichia coli/efeitos dos fármacos , Proteínas de Ligação às Penicilinas/antagonistas & inibidores , Andinocilina/química , Antibacterianos/química , Escherichia coli/metabolismo , Proteínas de Escherichia coli/metabolismo , Homeostase/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Proteínas de Ligação às Penicilinas/metabolismoRESUMO
The development of new antibiotics to treat infections caused by drug-resistant Gram-negative pathogens is of paramount importance as antibiotic resistance continues to increase worldwide1. Here we describe a strategy for the rational design of diazabicyclooctane inhibitors of penicillin-binding proteins from Gram-negative bacteria to overcome multiple mechanisms of resistance, including ß-lactamase enzymes, stringent response and outer membrane permeation. Diazabicyclooctane inhibitors retain activity in the presence of ß-lactamases, the primary resistance mechanism associated with ß-lactam therapy in Gram-negative bacteria2,3. Although the target spectrum of an initial lead was successfully re-engineered to gain in vivo efficacy, its ability to permeate across bacterial outer membranes was insufficient for further development. Notably, the features that enhanced target potency were found to preclude compound uptake. An improved optimization strategy leveraged porin permeation properties concomitant with biochemical potency in the lead-optimization stage. This resulted in ETX0462, which has potent in vitro and in vivo activity against Pseudomonas aeruginosa plus all other Gram-negative ESKAPE pathogens, Stenotrophomonas maltophilia and biothreat pathogens. These attributes, along with a favourable preclinical safety profile, hold promise for the successful clinical development of the first novel Gram-negative chemotype to treat life-threatening antibiotic-resistant infections in more than 25 years.
Assuntos
Antibacterianos/farmacologia , Desenho de Fármacos , Farmacorresistência Bacteriana Múltipla , Bactérias Gram-Negativas/efeitos dos fármacos , Animais , Antibacterianos/química , Compostos Aza/química , Compostos Aza/farmacologia , Ciclo-Octanos/química , Ciclo-Octanos/farmacologia , Feminino , Camundongos , Camundongos Endogâmicos BALB C , Estrutura Molecular , Proteínas de Ligação às Penicilinas/antagonistas & inibidores , Pseudomonas aeruginosa/efeitos dos fármacos , beta-LactamasesRESUMO
The effectiveness of ß-lactam antibiotics is increasingly compromised by ß-lactamases. Boron-containing inhibitors are potent serine-ß-lactamase inhibitors, but the interactions of boron-based compounds with the penicillin-binding protein (PBP) ß-lactam targets have not been extensively studied. We used high-throughput X-ray crystallography to explore reactions of a boron-containing fragment set with the Pseudomonas aeruginosa PBP3 (PaPBP3). Multiple crystal structures reveal that boronic acids react with PBPs to give tricovalently linked complexes bonded to Ser294, Ser349, and Lys484 of PaPBP3; benzoxaboroles react with PaPBP3 via reaction with two nucleophilic serines (Ser294 and Ser349) to give dicovalently linked complexes; and vaborbactam reacts to give a monocovalently linked complex. Modifications of the benzoxaborole scaffold resulted in a moderately potent inhibition of PaPBP3, though no antibacterial activity was observed. Overall, the results further evidence the potential for the development of new classes of boron-based antibiotics, which are not compromised by ß-lactamase-driven resistance.
Assuntos
Antibacterianos/farmacologia , Compostos de Boro/farmacologia , Ensaios de Triagem em Larga Escala , Proteínas de Ligação às Penicilinas/antagonistas & inibidores , Pseudomonas aeruginosa/efeitos dos fármacos , Inibidores de beta-Lactamases/farmacologia , Antibacterianos/síntese química , Antibacterianos/química , Sítios de Ligação/efeitos dos fármacos , Compostos de Boro/síntese química , Compostos de Boro/química , Cristalografia por Raios X , Relação Dose-Resposta a Droga , Testes de Sensibilidade Microbiana , Modelos Moleculares , Estrutura Molecular , Proteínas de Ligação às Penicilinas/metabolismo , Relação Estrutura-Atividade , Inibidores de beta-Lactamases/síntese química , Inibidores de beta-Lactamases/química , beta-LactamasesRESUMO
Penicillin-binding protein 2a (PBP2a) is an essential protein involved in the resistance to ß-lactam antibiotics acquired by methicillin-resistant Staphylococcus aureus (MRSA) and is a potential antibacterial target. In the current study, we employed a strategy that combined virtual screening with biological evaluation to discover novel inhibitors of PBP2a. In this investigation, a hybrid virtual screening method, consisting of drug-likeness evaluation (Lipinski's Rule of Five and ADMET) and rigid (LibDock) and semi-flexible (CDOCKER) docking-based virtual screenings, was used for retrieving novel PBP2a inhibitors from commercially available chemical databases. 11 compounds were selected from the final hits and subsequently shifted to experimental studies. Among them, Hit 2, Hit 3, and Hit 10 exhibited excellent anti-MRSA ATCC 33591 activity and weak toxicity in vitro. The affinity of the three compounds to bind to PBP2a was further confirmed by surface plasmon resonance (SPR) experiments and molecular dynamics (MD) simulation. An inter-complex interaction study showed that all hit compounds adapted well to the allosteric site of the PBP2a protein. In addition, Hit 2 (with best binding affinity to PBP2a, KD = 1.29 × 10-7 M) significantly inhibits proliferation of MRSA clinical isolates. Together, the 3 hit compounds, especially Hit 2, may be potential non-ß-lactam antibiotics against MRSA and the work will provide clues for the future development of specific compounds that block the interaction of PBP2a with their targets.
Assuntos
Antibacterianos/farmacologia , Proteínas de Bactérias/antagonistas & inibidores , Descoberta de Drogas , Proteínas de Ligação às Penicilinas/antagonistas & inibidores , Staphylococcus aureus/efeitos dos fármacos , Antibacterianos/síntese química , Antibacterianos/química , Proteínas de Bactérias/metabolismo , Relação Dose-Resposta a Droga , Avaliação Pré-Clínica de Medicamentos , Testes de Sensibilidade Microbiana , Simulação de Dinâmica Molecular , Estrutura Molecular , Proteínas de Ligação às Penicilinas/metabolismo , Staphylococcus aureus/metabolismo , Relação Estrutura-AtividadeRESUMO
Multidrug-resistant (MDR) pathogens pose a significant public health threat. A major mechanism of resistance expressed by MDR pathogens is ß-lactamase-mediated degradation of ß-lactam antibiotics. The diazabicyclooctane (DBO) compounds zidebactam and WCK 5153, recognized as ß-lactam "enhancers" due to inhibition of Pseudomonas aeruginosa penicillin-binding protein 2 (PBP2), are also class A and C ß-lactamase inhibitors. To structurally probe their mode of PBP2 inhibition as well as investigate why P. aeruginosa PBP2 is less susceptible to inhibition by ß-lactam antibiotics compared to the Escherichia coli PBP2, we determined the crystal structure of P. aeruginosa PBP2 in complex with WCK 5153. WCK 5153 forms an inhibitory covalent bond with the catalytic S327 of PBP2. The structure suggests a significant role for the diacylhydrazide moiety of WCK 5153 in interacting with the aspartate in the S-X-N/D PBP motif. Modeling of zidebactam in the active site of PBP2 reveals a similar binding mode. Both DBOs increase the melting temperature of PBP2, affirming their stabilizing interactions. To aid in the design of DBOs that can inhibit multiple PBPs, the ability of three DBOs to interact with P. aeruginosa PBP3 was explored crystallographically. Even though the DBOs show covalent binding to PBP3, they destabilized PBP3. Overall, the studies provide insights into zidebactam and WCK 5153 inhibition of PBP2 compared to their inhibition of PBP3 and the evolutionarily related KPC-2 ß-lactamase. These molecular insights into the dual-target DBOs advance our knowledge regarding further DBO optimization efforts to develop novel potent ß-lactamase-resistant, non-ß-lactam PBP inhibitors.IMPORTANCE Antibiotic resistance is a significant clinical problem. Developing novel antibiotics that overcome known resistance mechanisms is highly desired. Diazabicyclooctane inhibitors such as zidebactam possess this potential as they readily inactivate penicillin-binding proteins, yet cannot be degraded by ß-lactamases. In this study, we characterized the inhibition by diazabicyclooctanes of penicillin-binding proteins PBP2 and PBP3 from Pseudomonas aeruginosa using protein crystallography and biophysical analyses. These structures and analyses help define the antibiotic properties of these inhibitors, explain the decreased susceptibility of P. aeruginosa PBP2 to be inhibited by ß-lactam antibiotics, and provide insights that could be used for further antibiotic development.
Assuntos
Antibacterianos/farmacologia , Compostos Azabicíclicos/farmacologia , Compostos Bicíclicos com Pontes/farmacologia , Ciclo-Octanos/farmacologia , Octanos/farmacologia , Proteínas de Ligação às Penicilinas/química , Proteínas de Ligação às Penicilinas/metabolismo , Piperidinas/farmacologia , Pseudomonas aeruginosa/química , Pseudomonas aeruginosa/efeitos dos fármacos , Antibacterianos/metabolismo , Compostos Azabicíclicos/metabolismo , Compostos Bicíclicos com Pontes/metabolismo , Cristalização , Ciclo-Octanos/metabolismo , Testes de Sensibilidade Microbiana , Octanos/metabolismo , Proteínas de Ligação às Penicilinas/antagonistas & inibidores , Piperidinas/metabolismo , Ligação Proteica , Pseudomonas aeruginosa/enzimologia , Pseudomonas aeruginosa/genética , Inibidores de beta-Lactamases/farmacologiaRESUMO
Antimicrobial resistance is an imminent threat worldwide. Methicillin-resistant Staphylococcus aureus (MRSA) is one of the "superbug" family, manifesting resistance through the production of a penicillin binding protein, PBP2a, an enzyme that provides its transpeptidase activity to allow cell wall biosynthesis. PBP2a's low affinity to most ß-lactams, confers resistance to MRSA against numerous members of this class of antibiotics. An Achilles' heel of MRSA, PBP2a represents a substantial target to design novel antibiotics to tackle MRSA threat via inhibition of the bacterial cell wall biosynthesis. In this review we bring into focus the PBP2a enzyme and examine the various aspects related to its role in conferring resistance to MRSA strains. Moreover, we discuss several antibiotics and antimicrobial agents designed to target PBP2a and their therapeutic potential to meet such a grave threat. In conclusion, we consider future perspectives for targeting MRSA infections.
Assuntos
Antibacterianos/farmacologia , Proteínas de Bactérias/antagonistas & inibidores , Inibidores Enzimáticos/farmacologia , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Proteínas de Ligação às Penicilinas/antagonistas & inibidores , Antibacterianos/química , Proteínas de Bactérias/metabolismo , Química Farmacêutica , Inibidores Enzimáticos/química , Staphylococcus aureus Resistente à Meticilina/enzimologia , Testes de Sensibilidade Microbiana , Proteínas de Ligação às Penicilinas/metabolismoRESUMO
Treatment of multidrug-resistant Gram-negative bacterial pathogens represents a critical clinical need. Here, we report a novel γ-lactam pyrazolidinone that targets penicillin-binding proteins (PBPs) and incorporates a siderophore moiety to facilitate uptake into the periplasm. The MIC values of γ-lactam YU253434, 1, are reported along with the finding that 1 is resistant to hydrolysis by all four classes of ß-lactamases. The druglike characteristics and mouse PK data are described along with the X-ray crystal structure of 1 binding to its target PBP3.
Assuntos
Antibacterianos/farmacologia , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Lactamas/química , Sideróforos/química , Animais , Antibacterianos/síntese química , Antibacterianos/metabolismo , Antibacterianos/farmacocinética , Proteínas de Bactérias/antagonistas & inibidores , Proteínas de Bactérias/metabolismo , Sítios de Ligação , Domínio Catalítico , Cristalografia por Raios X , Bactérias Gram-Negativas/efeitos dos fármacos , Meia-Vida , Lactamas/metabolismo , Lactamas/farmacocinética , Lactamas/farmacologia , Camundongos , Testes de Sensibilidade Microbiana , Simulação de Acoplamento Molecular , Proteínas de Ligação às Penicilinas/antagonistas & inibidores , Proteínas de Ligação às Penicilinas/metabolismo , Pseudomonas aeruginosa/metabolismo , Sideróforos/metabolismoRESUMO
The enzymes involved in bacterial cell wall synthesis are established antibiotic targets, and continue to be a central focus for antibiotic development. Bacterial penicillin-binding proteins (and, in some bacteria, l,d-transpeptidases) form essential peptide cross-links in the cell wall. Although the ß-lactam class of antibiotics target these enzymes, bacterial resistance threatens their clinical use, and there is an urgent unmet need for new antibiotics. However, the search for new antibiotics targeting the bacterial cell wall is hindered by a number of obstacles associated with screening the enzymes involved in peptidoglycan synthesis. This review describes recent approaches for measuring the activity and inhibition of penicillin-binding proteins and l,d-transpeptidases, highlighting strategies that are poised to serve as valuable tools for high-throughput screening of transpeptidase inhibitors, supporting the development of new antibiotics.
Assuntos
Antibacterianos/farmacologia , Bactérias/efeitos dos fármacos , Parede Celular/efeitos dos fármacos , Descoberta de Drogas , Peptidil Transferases/antagonistas & inibidores , Antibacterianos/química , Bactérias/metabolismo , Parede Celular/metabolismo , Testes de Sensibilidade Microbiana , Estrutura Molecular , Proteínas de Ligação às Penicilinas/antagonistas & inibidores , Proteínas de Ligação às Penicilinas/metabolismo , Peptidil Transferases/metabolismoRESUMO
Selective chemical probes enable individual investigation of penicillin-binding proteins (PBPs) and provide critical information about their enzymatic activity with spatial and temporal resolution. To identify scaffolds for novel probes to study peptidoglycan biosynthesis in Bacillus subtilis, we evaluated the PBP inhibition profiles of 21 ß-lactam antibiotics from different structural subclasses using a fluorescence-based assay. Most compounds readily labeled PBP1, PBP2a, PBP2b, or PBP4. Almost all penicillin scaffolds were coselective for all or combinations of PBP2a, 2b, and 4. Cephalosporins, on the other hand, possessed the lowest IC50 values for PBP1 alone or along with PBP4 (ceftriaxone, cefoxitin) and 2b (cefotaxime) or 2a, 2b, and 4 (cephalothin). Overall, five selective inhibitors for PBP1 (aztreonam, faropenem, piperacillin, cefuroxime, and cefsulodin), one selective inhibitor for PBP5 (6-aminopenicillanic acid), and various coselective inhibitors for other PBPs in B. subtilis were discovered. Surprisingly, carbapenems strongly inhibited PBP3, formerly shown to have low affinity for ß-lactams and speculated to be involved in ß-lactam resistance in B. subtilis. To investigate the specific roles of PBP3, we developed activity-based probes based on the meropenem core and utilized them to monitor the activity of PBP3 in living cells. We showed that PBP3 activity localizes as patches in single cells and concentrates as a ring at the septum and the division site during the cell growth cycle. Our activity-based approach enabled spatial resolution of the transpeptidation activity of individual PBPs in this model microorganism, which was not possible with previous chemical and biological approaches.
Assuntos
Antibacterianos/química , Bacillus subtilis/enzimologia , Inibidores Enzimáticos/química , Proteínas de Ligação às Penicilinas/antagonistas & inibidores , beta-Lactamas/química , Acetilglucosamina/metabolismo , Antibacterianos/farmacologia , Bacillus subtilis/metabolismo , Divisão Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Avaliação Pré-Clínica de Medicamentos , Ativação Enzimática , Inibidores Enzimáticos/farmacologia , Corantes Fluorescentes/química , Glicosilação , Humanos , Iluminação , Ácidos Murâmicos/metabolismo , Imagem Óptica , Relação Estrutura-Atividade , beta-Lactamas/farmacologiaRESUMO
Ceftobiprole is an advanced-generation broad-spectrum cephalosporin antibiotic with potent and rapid bactericidal activity against Gram-positive pathogens, including methicillin-resistant Staphylococcus aureus, as well as susceptible Gram-negative pathogens, including Pseudomonas sp. pathogens. In the case of Pseudomonas aeruginosa, ceftobiprole acts by inhibiting P. aeruginosa penicillin-binding protein 3 (PBP3). Structural studies were pursued to elucidate the molecular details of this PBP inhibition. The crystal structure of the His-tagged PBP3-ceftobiprole complex revealed a covalent bond between the ligand and the catalytic residue S294. Ceftobiprole binding leads to large active site changes near binding sites for the pyrrolidinone and pyrrolidine rings. The S528 to L536 region adopts a conformation previously not observed in PBP3, including partial unwinding of the α11 helix. These molecular insights can lead to a deeper understanding of ß-lactam-PBP interactions that result in major changes in protein structure, as well as suggesting how to fine-tune current inhibitors and to develop novel inhibitors of this PBP.
Assuntos
Antibacterianos/farmacologia , Cefalosporinas/metabolismo , Proteínas de Ligação às Penicilinas/antagonistas & inibidores , Proteínas de Ligação às Penicilinas/metabolismo , Pseudomonas aeruginosa/efeitos dos fármacos , Antibacterianos/metabolismo , Sítios de Ligação/fisiologia , Domínio Catalítico/efeitos dos fármacos , Cefalosporinas/farmacologia , Cristalografia por Raios X , Humanos , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Conformação Molecular , Ligação ProteicaRESUMO
Because of ß-lactamase-mediated resistance, ß-lactam antibiotics were long considered ineffective drugs for tuberculosis (TB) treatment. However, some ß-lactams, including meropenem and faropenem, are being re-evaluated in patients infected with TB. Penicillin-binding protein (PBP) 3, or ftsI, is an essential transpeptidase in Mycobacterium tuberculosis (Mtb) required for cell division, and thus it is an important drug target. Structures of apo MtbPBP3 and of complexes with five ß-lactams, including meropenem and faropenem, reveal how they cause inactivation via formation of hydrolytically stable acyl-enzyme complexes. The structures reveal unique features of the antibiotic interactions, both in terms of differences in their binding to MtbPBP3 and in comparison with structures of other PBPs and serine ß-lactamases, including the tautomerization status of the carbapenem-derived acyl-enzyme complexes. The results suggest that rather than hoping PBP inhibitors developed for other infections will work against TB, work should focus on developing PBP inhibitors specialized for treating TB. SIGNIFICANCE STATEMENT: The structures of Mycobacterium tuberculosis penicillin-binding protein 3, an essential protein in M. tuberculosis, in complex with a number of widely used ß-lactam antibiotics (e.g., meropenem, aztreonam, and amoxicillin) were solved. These data provide new insights for next-generation rational approaches to design tuberculosis (TB)-specific ß-lactam or nonlactam antibiotics. This manuscript is a seminal article in the field of anti-TB drug discovery and suitable for the broad readership.
Assuntos
Antibacterianos/química , Mycobacterium tuberculosis/fisiologia , Proteínas de Ligação às Penicilinas/ultraestrutura , Resistência beta-Lactâmica , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Cristalografia por Raios X , Desenho de Fármacos , Meropeném/química , Meropeném/farmacologia , Testes de Sensibilidade Microbiana , Modelos Moleculares , Mycobacterium tuberculosis/efeitos dos fármacos , Proteínas de Ligação às Penicilinas/antagonistas & inibidores , Proteínas de Ligação às Penicilinas/metabolismo , Ligação Proteica , Domínios Proteicos , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/ultraestrutura , Tuberculose/tratamento farmacológico , Tuberculose/microbiologia , beta-Lactamas/química , beta-Lactamas/farmacologiaRESUMO
Background PBP2a is a type of penicillin-binding proteins (PBPs) that cause resistivity in methicillin-resistant Staphylococcus aureus (MRSA) from ß-lactam antibiotics. MRSA susceptible with cefttobiprole (fifth generation of cephalosporin as an anti-MRSA agent) which inhibits PBP2a and stops its growth. Contrary to its efficacy, ceftobiprole causes taste disturbance more than any other cephalosporins; furthermore, its mechanism is unknown. This study aims to explore an in silico study of a natural compound, which serves as a potential alternative to overcome MRSA with minimum adverse side effects. Methods A molecular docking study was performed using Molegro Virtual Docker version 5.5. Brazilin and proto-sappanins A-E are phytochemical compounds contained in sappan wood extract and are docked into the binding site of PBP2a (Protein Data Bank: ID 4DKI). Results Brazilin and proto-sappanins A-E have some interaction with Ser 403 amino acid residue which is an important interaction to inhibit PBP2a protein. The result of the molecular docking study showed that the MolDock score of proto-sappanins D and E is lower than that of methicillin but higher than that of its native ligand (ceftobiprole). Conclusions The results of this study suggest that proto-sappanins D and E have an excellent potential activity as an alternative to ceftobiprole in limiting MRSA growth through PBP2A enzyme inhibition.
Assuntos
Proteínas de Bactérias/antagonistas & inibidores , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Simulação de Acoplamento Molecular , Proteínas de Ligação às Penicilinas/antagonistas & inibidores , Compostos Fitoquímicos/farmacologia , Extratos Vegetais/farmacologia , Infecções Estafilocócicas/tratamento farmacológico , Madeira/química , Antibacterianos/farmacologia , Simulação por Computador , Humanos , Staphylococcus aureus Resistente à Meticilina/enzimologia , Infecções Estafilocócicas/microbiologia , Infecções Estafilocócicas/patologiaRESUMO
Protein-protein interactions are key in mycobacterial physiology, notably during the biosynthesis of the very peculiar mycobacterial cell wall. In this paper, we demonstrate that MSMEG_1285 interacts with PonA1, a bifunctional penicillin-binding protein involved in peptidoglycan biosynthesis. Deletion of MSMEG_1285 enhances Mycobacterium smegmatis resistance to penicillin antibiotics, a phenotype that is exacerbated by the additional deletion of hbhA. This also led to a substantial decrease in the amounts of porins in the cell wall, which are necessary for the import of small and hydrophilic ß-lactams. Deletion of both MSMEG_1285 and hbhA provoked an over-representation of several enzymes involved in peptidoglycan degradation. Thus, we propose that MSMEG_1285 is part of a protein scaffold, which also involves PonA1 and HbhA, and that it is responsible for the tight regulation of peptidoglycan hydrolysis. This study provides a better understanding of the mycobacterial physiology, which is an essential step for strengthening the action of drugs that specifically target peptidoglycan biosynthesis.
Assuntos
Antibacterianos/farmacologia , Proteínas de Bactérias/antagonistas & inibidores , Parede Celular/efeitos dos fármacos , Mycobacterium/efeitos dos fármacos , Proteínas de Ligação às Penicilinas/antagonistas & inibidores , Penicilinas/farmacologia , Proteínas de Bactérias/metabolismo , Parede Celular/metabolismo , Hidrólise , Testes de Sensibilidade Microbiana , Mycobacterium/metabolismo , Proteínas de Ligação às Penicilinas/metabolismo , Peptidoglicano/metabolismoRESUMO
Monocyclic ß-lactams revive the research field on antibiotics, which are threatened by the emergence of resistant bacteria. A six-step synthetic route was developed, providing easy access to new 3-amino-1-carboxymethyl-4-phenyl-ß-lactams, of which the penicillin-binding protein (PBP) inhibitory potency was demonstrated biochemically.
Assuntos
Aminoácidos/farmacologia , Antibacterianos/farmacologia , Dipeptídeos/farmacologia , Iminas/farmacologia , Lactamas/farmacologia , Proteínas de Ligação às Penicilinas/antagonistas & inibidores , Aminoácidos/química , Antibacterianos/síntese química , Antibacterianos/química , Dipeptídeos/síntese química , Dipeptídeos/química , Iminas/química , Lactamas/síntese química , Lactamas/química , Estrutura Molecular , Proteínas de Ligação às Penicilinas/metabolismoRESUMO
Methicillin-resistant Staphylococcus aureus (MRSA) is a multidrug-resistant bacterium responsible for serious nosocomial and community-acquired infections worldwide. Since few antibiotics are effective for treating MRSA infections, the development of new therapies is of great importance. Previous studies demonstrated that PBP2a is a target that generates protective antibodies against MRSA. A murine monoclonal antibody (MAb) that recognizes PBP2a from MRSA strains was previously isolated and characterized. In this report, we evaluated the biodistribution of this MAb in blood and tissues, as well as the extent of protection conferred using prophylactic and therapeutic assays compared to vancomycin treatment. Biodistribution was evaluated 12-96 h after MAb administration. It predominantly remained in the serum, but it was also detectable in the kidneys, lungs, and spleen at low concentrations (about 4.5% in the kidneys, 1.9% in the lungs, and 0.7% the spleen) at all observed timepoints. Prophylactic studies in a murine model demonstrated a significant bacterial load reduction in the kidneys of the groups treated with either with IgG (greater than 3 logs) or F(ab')2 (98%) when compared to that of the control groups (untreated). Mice were challenged with a lethal dose, and the survival rate was higher in the treated mice. Treatment with the MAb resulted in a bacterial load reduction in the kidneys similar to that of mice treated with vancomycin, and a MAb/vancomycin combination therapy was also effective. These results demonstrate that an anti-PBP2a MAb may be a promising therapeutic for treating MRSA infections.
Assuntos
Antibacterianos/farmacologia , Anticorpos Antibacterianos/farmacologia , Anticorpos Monoclonais/farmacologia , Proteínas de Bactérias/antagonistas & inibidores , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Proteínas de Ligação às Penicilinas/antagonistas & inibidores , Substâncias Protetoras/farmacologia , Infecções Estafilocócicas/prevenção & controle , Animais , Carga Bacteriana , Proteínas de Bactérias/imunologia , Feminino , Staphylococcus aureus Resistente à Meticilina/crescimento & desenvolvimento , Camundongos , Camundongos Endogâmicos BALB C , Testes de Sensibilidade Microbiana , Proteínas de Ligação às Penicilinas/imunologia , Infecções Estafilocócicas/microbiologia , Vancomicina/farmacologiaRESUMO
Ceftobiprole, a novel last generation parenteral cephalosporin, has an extended spectrum of activity, notably against methicillin-resistant Staphylococcus aureus (MRSA), ampicillin-susceptible enterococci, penicillin-resistant pneumococci, Enterobacterales and susceptible Pseudomonas aeruginosa. It exerts an inhibitory action on essential peptidoglycan transpeptidases, interfering with cell wall synthesis. The inhibitory action of ceftobiprole through binding to abnormal PBPs like PBP2a in methicillin-resistant staphylococci and PBP2b and PBP2x in the case of ß-lactam-resistant pneumococci, ultimately leads to rapid bacterial cell death. In the case of Enterobacterales, ceftobiprole retains activity against narrow spectrum ß-lactamases but is hydrolysed by their extended-spectrum counterparts, overexpressed Amp C, and carbapenemases. It is also affected by certain efflux pumps from P. aeruginosa. For anaerobic bacteria, ceftobiprole is active against Gram-positive Clostridioides difficile and Peptococcus spp. and Gram-negative Fusobacterium nucleatum but not against Bacteroides group or other anaerobic Gram-negatives. In in vitro studies, a low propensity to select for resistant subpopulations has been demonstrated. Currently, ceftobiprole is approved for the treatment of community-acquired pneumonia and hospital-acquired pneumonia with the exception of ventilator-associated pneumonia. Ceftobiprole's place in therapy appears to lie mainly in its combined activity against Gram-positive organisms, such as S. aureus and S. pneumoniae alongside that against Gram-negative organisms such as P. aeruginosa.
Assuntos
Antibacterianos/farmacologia , Cefalosporinas/farmacologia , Aminoaciltransferases/antagonistas & inibidores , Antibacterianos/metabolismo , Cefalosporinas/metabolismo , Infecções Comunitárias Adquiridas/tratamento farmacológico , Endopeptidases/efeitos dos fármacos , Enterobacteriaceae/efeitos dos fármacos , Enterococcus/efeitos dos fármacos , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Positivas/efeitos dos fármacos , Bactérias Gram-Positivas/metabolismo , Humanos , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Resistência às Penicilinas , Proteínas de Ligação às Penicilinas/antagonistas & inibidores , Pneumonia Associada à Ventilação Mecânica/tratamento farmacológico , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/metabolismo , Streptococcus pneumoniae/efeitos dos fármacos , Inibidores de beta-Lactamases/farmacologiaRESUMO
The aim of the study was to determine the efficacy of dual ß-lactam combination treatments derived from eight approved drugs against Galleria mellonella larvae infected with MDR strains of P. aeruginosa. Carbapenem-resistant P. aeruginosa NCTC 13437 and an unrelated clinical isolate were used to infect G. mellonella larvae and the efficacy of twenty-eight dual ß-lactam combination therapies were compared to their constituent monotherapies. For the most potent combinations identified, penicillin-binding protein (PBP) inhibition profiles were measured and compared with each constituent antibiotic. Five of the dual ß-lactam combinations resulted in greater than 70% survival of infected G. mellonella. Two combinations showed potent, enhanced efficacy versus both strains - ceftazidime + meropenem and aztreonam + meropenem. Comparison of PBP inhibition profiles revealed that the enhanced efficacy of these two dual ß-lactam combinations could not be explained by more potent inhibition of PBPs or inhibition of a broader range of PBPs. A possible contribution to the enhanced efficacy of the combinations could be stimulation of innate immunity via increased haemocyte numbers compared to their constituent monotherapies. Combinations of ß-lactam antibiotics show promise in overcoming MDR P. aeruginosa and are worthy of additional study and development.