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1.
J Med Chem ; 67(13): 10774-10782, 2024 Jul 11.
Artigo em Inglês | MEDLINE | ID: mdl-38900970

RESUMO

Antibiotic resistance is an urgent threat to global health, with the decreasing efficacy of conventional drugs underscoring the urgency for innovative therapeutic strategies. Antimicrobial peptides present as promising alternatives to conventional antibiotics. Gramicidin S is one such naturally occurring antimicrobial peptide that is effective against Staphylococcus aureus, with a minimum inhibitory concentration (MIC) of 4 µg/mL (3.6 µM). Despite this potent activity, its significant hemolytic toxicity restricts its clinical use to topical applications. Herein, we present rational modifications to the key ß-strand and ß-turn regions of gramicidin S to concurrently mitigate hemolytic effects, while maintaining potency. Critically, peptide 9 displayed negligible hemolytic toxicity, while possessing significant antibacterial potency against a panel of methicillin-sensitive and methicillin-resistant S. aureus clinical isolates (MIC of 8 µg/mL, 7.2 µM). Given the substantial antibacterial activity and near absence of cytotoxicity, 9 presents as a potential candidate for systemic administration in the treatment of S. aureus bacteremia/sepsis.


Assuntos
Antibacterianos , Gramicidina , Hemólise , Testes de Sensibilidade Microbiana , Antibacterianos/farmacologia , Antibacterianos/química , Antibacterianos/síntese química , Hemólise/efeitos dos fármacos , Gramicidina/farmacologia , Gramicidina/análogos & derivados , Humanos , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Relação Estrutura-Atividade , Eritrócitos/efeitos dos fármacos , Animais
2.
Nat Commun ; 11(1): 4935, 2020 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-33004797

RESUMO

Gramicidin A (1) is a peptide antibiotic that disrupts the transmembrane ion concentration gradient by forming an ion channel in a lipid bilayer. Although long used clinically, it is limited to topical application because of its strong hemolytic activity and mammalian cytotoxicity, likely arising from the common ion transport mechanism. Here we report an integrated high-throughput strategy for discovering analogues of 1 with altered biological activity profiles. The 4096 analogue structures are designed to maintain the charge-neutral, hydrophobic, and channel forming properties of 1. Synthesis of the analogues, tandem mass spectrometry sequencing, and 3 microscale screenings enable us to identify 10 representative analogues. Re-synthesis and detailed functional evaluations find that all 10 analogues share a similar ion channel function, but have different cytotoxic, hemolytic, and antibacterial activities. Our large-scale structure-activity relationship studies reveal the feasibility of developing analogues of 1 that selectively induce toxicity toward target organisms.


Assuntos
Antibacterianos/farmacologia , Descoberta de Drogas/métodos , Gramicidina/análogos & derivados , Ensaios de Triagem em Larga Escala/métodos , Animais , Antibacterianos/química , Linhagem Celular Tumoral , Química Farmacêutica , Eritrócitos , Estudos de Viabilidade , Bactérias Gram-Positivas/efeitos dos fármacos , Gramicidina/química , Gramicidina/farmacologia , Hemólise/efeitos dos fármacos , Concentração Inibidora 50 , Camundongos , Testes de Sensibilidade Microbiana , Estrutura Molecular , Coelhos , Relação Estrutura-Atividade , Espectrometria de Massas em Tandem
3.
ChemMedChem ; 15(12): 1089-1100, 2020 06 17.
Artigo em Inglês | MEDLINE | ID: mdl-32233075

RESUMO

Gramicidin S (GS), one of the most widely investigated antimicrobial peptides (AMPs), is known for its robust antimicrobial activity. However, it is restricted to topical application due to undesired hemolytic activity. With the aim of obtaining nontoxic GS analogues, we describe herein a molecular approach in which the native GS ß-turn region is replaced by synthetic ß,γ-diamino acids (ß,γ-DiAAs). Four ß,γ-DiAA diastereomers were employed to mimic the ß-turn structure to afford GS analogues GS3-6, which exhibit diminished hemolytic activity. A comparative structural study demonstrates that the (ßR,γS)-DiAA is the most-stable ß-turn mimic. To further improve the therapeutic index (e. g., high antibacterial activity and low hemolytic activity) and to extend the molecular diversity, GS5 and GS6 were used as structural scaffolds to introduce additional hydrophobic or hydrophilic groups. We show that GS6K, GS6F and GS display comparable antibacterial activity, and GS6K and GS6F have significantly decreased toxicity. Moreover, antibacterial mechanism studies suggest that GS6K kills bacteria mainly through the disruption of the membrane.


Assuntos
Antibacterianos/farmacologia , Gramicidina/análogos & derivados , Gramicidina/farmacologia , Antibacterianos/síntese química , Antibacterianos/toxicidade , Bactérias/efeitos dos fármacos , Linhagem Celular , Membrana Celular/efeitos dos fármacos , Eritrócitos/efeitos dos fármacos , Gramicidina/toxicidade , Hemólise/efeitos dos fármacos , Humanos , Testes de Sensibilidade Microbiana , Estereoisomerismo
4.
Chem Pharm Bull (Tokyo) ; 68(2): 173-178, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32009085

RESUMO

An ion-channel-forming natural peptide, gramicidin A (1), exhibits potent antimicrobial activity against Gram-positive bacteria, although medical applications are limited to topical use due to its mammalian cytotoxicity. We recently reported that the artificial macrocyclic analogue 2 provides a promising starting point for developing new ion-channel-based systemic antibacterial agents because of its low mammalian cytotoxicity compared to that of the parent 1. To dissect the molecular factors involved in the species selectivity of 2, we evaluated the ion transport activities, phospholipid affinities, and conformational properties of 1 and 2 using various compositions of phospholipids. A combination of lipid dot blot assays and circular dichroism (CD) analysis with H+/Na+ exchange assays revealed that the higher H+/Na+ exchange activity of 2 than that of 1 in liposomes containing 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylcholine (POPC) or 1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-(1'-rac-glycerol) (POPG) is attributable to its higher affinity towards the phospholipids than that of 1. Notably, we also discovered that 2 showed weaker H+/Na+ exchange activity in liposomes containing 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylethanolamine (POPE). CD analysis of 2 in liposomes indicated that the weak H+/Na+ exchange activity is induced by disturbance of the ion-conducting ß6.3-helical conformation in the POPE-containing lipid bilayer. These results suggest that the POPE-induced attenuation of the ion-conducting activity of 2 contributes to the alleviation of undesirable mammalian cytotoxicity of 2 compared to that of 1.


Assuntos
Antibacterianos/química , Antibacterianos/farmacologia , Gramicidina/análogos & derivados , Gramicidina/farmacologia , Fosfolipídeos/metabolismo , Antibacterianos/toxicidade , Bactérias Gram-Positivas/efeitos dos fármacos , Infecções por Bactérias Gram-Positivas/tratamento farmacológico , Gramicidina/toxicidade , Humanos , Transporte de Íons/efeitos dos fármacos , Bicamadas Lipídicas/metabolismo , Compostos Macrocíclicos/química , Compostos Macrocíclicos/farmacologia
5.
Chembiochem ; 19(24): 2591-2597, 2018 12 18.
Artigo em Inglês | MEDLINE | ID: mdl-30324702

RESUMO

Gramicidin S is a naturally occurring antimicrobial cyclic peptide. Herein, we present a series of cyclic peptides based on gramicidin S that contain an azobenzene photoswitch to reversibly control secondary structure and, hence, antimicrobial activity. 1 H NMR spectroscopy and density functional theory calculations revealed a ß-sheet/ß-turn secondary structure for the cis configuration of each peptide, and an ill-defined conformation for all associated trans structures. The cis-enriched and trans-enriched photostationary states (PSSs) for peptides 1-3 were assayed against Staphylococcus aureus to reveal a clear relationship between well-defined secondary structure, amphiphilicity and optimal antimicrobial activity. Most notably, peptides 2 a and 2 b exhibited a fourfold difference in antimicrobial activity in the cis-enriched PSS over the trans-enriched equivalent. This photopharmacological approach allows antimicrobial activity to be regulated through photochemical control of the azobenzene photoswitch, thereby opening new avenues in the design and synthesis of future antibiotics.


Assuntos
Antibacterianos/farmacologia , Compostos Azo/farmacologia , Gramicidina/análogos & derivados , Gramicidina/farmacologia , Antibacterianos/síntese química , Antibacterianos/química , Antibacterianos/efeitos da radiação , Compostos Azo/síntese química , Compostos Azo/química , Compostos Azo/efeitos da radiação , Ciclização , Gramicidina/síntese química , Gramicidina/química , Isomerismo , Testes de Sensibilidade Microbiana , Modelos Moleculares , Staphylococcus aureus/efeitos dos fármacos , Raios Ultravioleta
6.
Anal Chem ; 90(3): 1635-1642, 2018 02 06.
Artigo em Inglês | MEDLINE | ID: mdl-29266927

RESUMO

Methods to detect low concentrations of small molecules are useful for a wide range of analytical problems including the development of clinical assays, the study of complex biological systems, and the detection of biological warfare agents. This paper describes a semisynthetic ion channel platform capable of detecting small molecule analytes with picomolar sensitivity. Our methodology exploits the transient nature of ion channels formed from gramicidin A (gA) nanopores and the frequency of observed single channel events as a function of concentration of free gA molecules that reversibly dimerize in a bilayer membrane. We initially use a protein (here, a monoclonal antibody) to sequester the ion channel activity of a C-terminally modified gA derivative. When a small molecule analyte is introduced to the electrical recording medium, it competitively binds to the protein and liberates the gA derivative, restoring its single ion channel activity. We found that monitoring the frequency of gA channel events makes it possible to detect picomolar concentrations of small molecule in solution. In part, due to the digital on/off nature of frequency-based analysis, this approach is 103 times more sensitive than measuring macroscopic membrane ion flux through gA channels as a basis for detection. This novel methodology, therefore, significantly improves the limit of detection of nanopore-based sensors for small molecule analytes, which has the potential for incorporation into miniaturized and low cost devices that could complement current established assays.


Assuntos
Técnicas Biossensoriais/métodos , Fluoresceínas/análise , Gramicidina/metabolismo , Canais Iônicos/metabolismo , Bicamadas Lipídicas/metabolismo , Anticorpos Monoclonais/imunologia , Fluoresceínas/síntese química , Fluoresceínas/química , Gramicidina/análogos & derivados , Gramicidina/síntese química , Haptenos/imunologia , Limite de Detecção , Bicamadas Lipídicas/química , Nanoporos
7.
Bioorg Med Chem ; 25(1): 261-268, 2017 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-27865644

RESUMO

The d-/l-peptide gramicidin A (gA) is well known as a pivotal ion channel model and shows a broad spectrum of bioactivities such as antibiosis, antimalarial activity, as well as hemolysis. We applied inter-chain disulfide bonds to constrain the conformational freedom of gA into parallel and antiparallel dimeric topologies. Albeit the constructs were not found to be monoconformational, CD- and IR-spectroscopic studies suggested that this strategy indeed restricted the conformational space of the d-/l-peptide construct, and that ß-helical secondary structures prevail. Correlative testing of gA dimers in antimicrobial, antimalarial, and ion conduction assays suggested that the tail-to-tail antiparallel single stranded ß6.3 helix dominantly mediates the bioactivity of gA. Other conformers are unlikely to contribute to these activities. From these investigations, only weakly ion conducting gA dimers were identified that retained nM antimalarial activity.


Assuntos
Antibacterianos/farmacologia , Antimaláricos/farmacologia , Dissulfetos/farmacologia , Gramicidina/análogos & derivados , Gramicidina/farmacologia , Antibacterianos/síntese química , Antimaláricos/síntese química , Dicroísmo Circular , Dimerização , Dissulfetos/síntese química , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Positivas/efeitos dos fármacos , Gramicidina/síntese química , Hemólise , Membranas Artificiais , Conformação Molecular , Permeabilidade , Plasmodium falciparum/efeitos dos fármacos
8.
ChemMedChem ; 11(6): 629-36, 2016 Mar 17.
Artigo em Inglês | MEDLINE | ID: mdl-26918268

RESUMO

Antimicrobial peptides (AMPs) have shown potential as alternatives to traditional antibiotics for fighting infections caused by antibiotic-resistant bacteria. One promising example of this is gramicidin A (gA). In its wild-type sequence, gA is active by permeating the plasma membrane of Gram-positive bacteria. However, gA is toxic to human red blood cells at similar concentrations to those required for it to exert its antimicrobial effects. Installing cationic side chains into gA has been shown to lower its hemolytic activity while maintaining the antimicrobial potency. In this study, we present the synthesis and the antibiotic activity of a new series of gA mutants that display cationic side chains. Specifically, by synthesizing alkylated lysine derivatives through reductive amination, we were able to create a broad selection of structures with varied activities towards Staphylococcus aureus and methicillin-resistant S. aureus (MRSA). Importantly, some of the new mutants were observed to have an unprecedented activity towards important Gram-negative pathogens, including Escherichia coli, Klebsiella pneumoniae and Psuedomonas aeruginosa.


Assuntos
Antibacterianos/farmacologia , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Positivas/efeitos dos fármacos , Gramicidina/análogos & derivados , Gramicidina/farmacologia , Lisina/análogos & derivados , Lisina/farmacologia , Antibacterianos/síntese química , Permeabilidade da Membrana Celular/efeitos dos fármacos , Gramicidina/síntese química , Hemólise , Lisina/síntese química , Relação Estrutura-Atividade
9.
Org Biomol Chem ; 13(24): 6789-802, 2015 Jun 28.
Artigo em Inglês | MEDLINE | ID: mdl-26008215

RESUMO

Gramicidin S (GS) is a cyclic cationic antimicrobial peptide (CAP) with a wide spectrum of antibiotic activities whose usage has been limited to topical applications owing to its cytotoxic side effects. We have synthesized tetrahydrofuran amino acid (Taa)-containing GS analogues, and we have carried out conformational analysis and explored their structure activity relationships by evaluating their antitubercular, antibacterial and cytotoxic properties. Two of these analogues showed impressive as well as selective activity against Mycobacterium tuberculosis (MTB) without toxicity towards mammalian Vero cells or human RBCs, and are promising as potential leads.


Assuntos
Antibacterianos/química , Antibacterianos/farmacologia , Furanos/química , Furanos/farmacologia , Gramicidina/análogos & derivados , Gramicidina/farmacologia , Animais , Antituberculosos/química , Antituberculosos/farmacologia , Bactérias/efeitos dos fármacos , Infecções Bacterianas/tratamento farmacológico , Infecções Bacterianas/microbiologia , Chlorocebus aethiops , Eritrócitos/efeitos dos fármacos , Humanos , Testes de Sensibilidade Microbiana , Simulação de Dinâmica Molecular , Mycobacterium tuberculosis/efeitos dos fármacos , Tuberculose/tratamento farmacológico , Tuberculose/microbiologia , Células Vero
10.
ChemMedChem ; 10(3): 540-5, 2015 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-25510221

RESUMO

A linear peptide, gramicidin A (GA), folds into a ß(6.3) -helix, functions as an ion channel in the cell membrane, and exerts antibacterial activity. Herein we describe the rational design, synthesis, and biological evaluation of lactam-bridged GA analogues. The GA analogue with a 27-membered macrolactam was found to adopt a stable ß(6.3) -helical conformation and exhibits higher ion-exchange activity than GA. Furthermore, this GA analogue retains the potent antibiotic activity of GA, but its hemolytic activity and toxicity toward mammalian cells are significantly lower than those of GA. This study thus dissociates the antibacterial and hemolytic/cytotoxic activities of GA, and charts a rational path forward for the development of new ion-channel-based antibiotics.


Assuntos
Antibacterianos/química , Antibacterianos/farmacologia , Gramicidina/análogos & derivados , Gramicidina/farmacologia , Peptídeos/química , Peptídeos/farmacologia , Antibacterianos/efeitos adversos , Bactérias/efeitos dos fármacos , Infecções Bacterianas/tratamento farmacológico , Linhagem Celular , Descoberta de Drogas , Gramicidina/efeitos adversos , Hemólise/efeitos dos fármacos , Humanos , Modelos Moleculares , Peptídeos/efeitos adversos , Estrutura Secundária de Proteína
11.
ChemMedChem ; 8(11): 1865-72, 2013 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-24023000

RESUMO

ß-Sheet antimicrobial peptides (AMPs) are well recognized as promising candidates for the treatment of multidrug-resistant bacterial infections. To dissociate antimicrobial activity and hemolytic effect of ß-sheet AMPs, we hypothesize that N-methylation of the intramolecular hydrogen bond(s)-forming amides could improve their specificities for microbial cells over human erythrocytes. We utilized a model ß-sheet antimicrobial peptide, gramicidin S (GS), to study the N-methylation effects on the antimicrobial and hemolytic activities. We synthesized twelve N-methylated GS analogues by replacement of residues at the ß-strand and ß-turn regions with N-methyl amino acids, and tested their antimicrobial and hemolytic activities. Our experiments showed that the HC50 values increased fivefold compared with that of GS, when the internal hydrogen-bonded leucine residue was methylated. Neither hemolytic effect nor antimicrobial activity changed when proline alone was replaced with N-methylalanine in the ß-turn region. However, analogues containing N-methylleucine at ß-strand and N-methylalanine at ß-turn regions exhibited a fourfold increase in selectivity index compared to GS. We also examined the conformation of these N-methylated GS analogues using (1)H NMR and circular dichroism (CD) spectroscopy in aqueous solution, and visualized the backbone structures and residue orientations using molecular dynamics simulations. The results show that N-methylation of the internal hydrogen bond-forming amide affected the conformation, backbone shape, and side chain orientation of GS.


Assuntos
Alanina/análogos & derivados , Anti-Infecciosos/síntese química , Anti-Infecciosos/farmacologia , Gramicidina/síntese química , Gramicidina/farmacologia , Alanina/química , Anti-Infecciosos/química , Bactérias/efeitos dos fármacos , Gramicidina/análogos & derivados , Hemólise/efeitos dos fármacos , Espectroscopia de Ressonância Magnética , Estrutura Secundária de Proteína
12.
Dalton Trans ; 42(6): 1973-8, 2013 Feb 14.
Artigo em Inglês | MEDLINE | ID: mdl-23235486

RESUMO

The cyclic peptide gramicidin S was used as a rigid template to provide novel peptide-based bisphosphine ligands for transition metal catalysis. Two bisphosphine-coordinated Rh(I) complexes allowed asymmetric hydrogenation with 10-52% ee and the corresponding Pd(II) analogues catalysed asymmetric allylic alkylation with 13-15% ee.


Assuntos
Gramicidina/análogos & derivados , Ligantes , Fosfinas/química , Alquilação , Catálise , Complexos de Coordenação/química , Gramicidina/síntese química , Hidrogenação , Paládio/química , Ródio/química
13.
Bioorg Med Chem ; 20(20): 6059-62, 2012 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-22989907

RESUMO

A series of gramicidin S derivatives 4-15 are presented that have four ornithine residues as polar protonated side chains and two central hydrophobic amino acids with unaltered turn regions. These peptides were screened against human erthrocytes and our standard panel of Gram negative- and Gram positive bacteria, including four MRSA strains. Based on the antibacterial- and hemolytic data, peptides 13 and 14 have an improved biological profile compared to the clinically applied topical antibiotic gramicidin S.


Assuntos
Antibacterianos/química , Gramicidina/análogos & derivados , Gramicidina/química , Antibacterianos/síntese química , Antibacterianos/farmacologia , Eritrócitos/efeitos dos fármacos , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Positivas/efeitos dos fármacos , Gramicidina/síntese química , Gramicidina/farmacologia , Hemólise , Humanos , Testes de Sensibilidade Microbiana , Peptídeos/síntese química , Peptídeos/química , Peptídeos/farmacologia
14.
Chem Pharm Bull (Tokyo) ; 60(9): 1134-8, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22976321

RESUMO

The substitution of each constituent amino acid residue of gramicidin S (GS), cyclo(-Val(1,1')-Orn(2,2')-Leu(3,3')-D-Phe(4,4')-Pro(5,5')-)(2) with Lys residue indicated that each side chain structure of the constituent amino acid residues affect largely the antibiotic activity and hemolytic activity of GS. Further, the substitution of D-Phe(4,4') and Pro(5,5') residues with basic amino acid residues as a Lys residue results the high antibiotic activity and the very low hemolytic activity. Thus, we have found novel positions on the scaffold of GS at D-Phe(4,4') and Pro(5,5') residues whose modification will significantly increase the therapeutic index.


Assuntos
Antibacterianos/química , Antibacterianos/farmacologia , Gramicidina/análogos & derivados , Gramicidina/farmacologia , Hemólise/efeitos dos fármacos , Animais , Antibacterianos/efeitos adversos , Bactérias/efeitos dos fármacos , Infecções Bacterianas/tratamento farmacológico , Eritrócitos/efeitos dos fármacos , Gramicidina/efeitos adversos , Ovinos
15.
PLoS One ; 7(7): e41919, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22911866

RESUMO

Limited uncoupling of oxidative phosphorylation could be beneficial for cells by preventing excessive generation of reactive oxygen species. Typical uncouplers are weak organic acids capable of permeating across membranes with a narrow gap between efficacy and toxicity. Aimed at designing a nontoxic uncoupler, the protonatable amino acid residue Glu was substituted for Val at the N-terminus of the pentadecapeptide gramicidin A (gA). The modified peptide [Glu1]gA exhibited high uncoupling activity in isolated mitochondria, in particular, abolishing membrane potential at the inner mitochondrial membrane with the same or even larger efficacy as gA. With mitochondria in cell culture, the depolarizing activity of [Glu1]gA was observed at concentrations by an order of magnitude lower than those of gA. On the contrary, [Glu1]gA was much less potent in forming proton channels in planar lipid bilayers than gA. Remarkably, at uncoupling concentrations, [Glu1]gA did not alter cell morphology and was nontoxic in MTT test, in contrast to gA showing high toxicity. The difference in the behavior of [Glu1]gA and gA in natural and artificial membranes could be ascribed to increased capability of [Glu1]gA to permeate through membranes and/or redistribute between different membranes. Based on the protective role of mild uncoupling, [Glu1]gA and some other proton-conducting gA analogues may be considered as prototypes of prospective therapeutic agents.


Assuntos
Substituição de Aminoácidos , Ácido Glutâmico/metabolismo , Gramicidina/análogos & derivados , Gramicidina/metabolismo , Mitocôndrias Hepáticas/metabolismo , Ionóforos de Próton/metabolismo , Desacopladores/metabolismo , Sequência de Aminoácidos , Animais , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Eletricidade , Gramicidina/química , Concentração de Íons de Hidrogênio/efeitos dos fármacos , Canais Iônicos/metabolismo , Rim/citologia , Bicamadas Lipídicas/metabolismo , Lipossomos/metabolismo , Masculino , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias Hepáticas/efeitos dos fármacos , Dados de Sequência Molecular , Peptídeos/química , Peptídeos/farmacologia , Fenazinas/metabolismo , Ratos
16.
Bioorg Med Chem Lett ; 22(1): 106-9, 2012 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-22172697

RESUMO

In the present study, novel eight GS derivatives having the octanoyl-(Lys)(n)- moieties, cyclo{-Val-Orn-Leu-d-Phe-Pro(4ß-NH-X)-Val-Orn-Leu-d-Phe-Pro-} {X=-H (1), and -(Lys)(n)-CO(CH(2))(6)CH(3)n=0 (2), 1 (3), 2 (4), and 3 (5)} and cyclo{-Val-Orn-Leu-d-Phe-Pro(4α-NH-X)-Val-Orn-Leu-d-Phe-Pro-} {X=-H (6), and -(Lys)(n)-CO(CH(2))(6)CH(3)n=1 (7), and 2 (8)} were synthesized. Among them, 4, 5 and 8 result the high antibiotic activity against both Gram-positive and Gram-negative microorganisms tested. In addition, 4 and 5 showed very low hemolytic activity compared with that of GS. Thus, the introduction of the excess amino groups and the fatty acyl moiety to the γ-NH(2) group of Pro(5) residue in GS molecule lowered the unwanted hemolytic activity and enhanced the desired antibiotic activity.


Assuntos
Química Farmacêutica/métodos , Gramicidina/análogos & derivados , Hemólise/efeitos dos fármacos , Animais , Antibacterianos/farmacologia , Células Sanguíneas/efeitos dos fármacos , Relação Dose-Resposta a Droga , Desenho de Fármacos , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Positivas/efeitos dos fármacos , Lisina/química , Testes de Sensibilidade Microbiana , Modelos Químicos , Ovinos , Relação Estrutura-Atividade
17.
Eur Biophys J ; 41(2): 129-38, 2012 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-22042158

RESUMO

Insertion of charged groups at the N-terminus of the gramicidin A (gA) amino acid sequence is considered to be fatal for peptide channel-forming activity because of hindrance to the head-to-head dimer formation. Here the induction of ionic conductivity in planar bilayer lipid membranes (BLM) was studied with gA analogs having lysine either in the first ([Lys1]gA) or the third ([Lys3]gA) position. If added to the bathing solution at neutral or acidic pH, these analogs, being protonated and thus positively charged, were unable to induce ionic current across BLM. By contrast, at pH 11 the induction of BLM conductivity was observed with both lysine-substituted analogs. Based on the dependence of the macroscopic current on the side of the peptide addition, sensitivity to calcium ions and susceptibility to sensitized photoinactivation, as well as on the single-channel properties of the analogs, we surmise that at alkaline pH [Lys1]gA formed channels with predominantly single-stranded structure of head-to-head helical dimers, whereas [Lys3]gA open channels had the double-stranded helical structure. CD spectra of the lysine-substituted analogs in liposomes were shown to be pH-dependent.


Assuntos
Substituição de Aminoácidos , Permeabilidade da Membrana Celular/efeitos dos fármacos , Membrana Celular/efeitos dos fármacos , Gramicidina/análogos & derivados , Gramicidina/farmacologia , Bicamadas Lipídicas/metabolismo , Lisina , Sequência de Aminoácidos , Membrana Celular/metabolismo , Condutividade Elétrica , Gramicidina/química , Concentração de Íons de Hidrogênio , Interações Hidrofóbicas e Hidrofílicas , Bicamadas Lipídicas/química , Lisina/química , Dados de Sequência Molecular
18.
PLoS One ; 7(12): e52839, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-23285199

RESUMO

Protein recruitment to specific membrane locations may be governed or facilitated by electrostatic attraction, which originates from a multivalent ligand. Here we explored the energetics of a model system in which this simple electrostatic recruitment mechanism failed. That is, basic poly-L-lysine binding to one leaflet of a planar lipid bilayer did not recruit the triply-charged peptide (O-Pyromellitylgramicidin). Clustering was only observed in cases where PLL was bound to both channel ends. Clustering was indicated (i) by the decreased diffusional PLL mobility D(PLL) and (ii) by an increased lifetime τ(PLL) of the clustered channels. In contrast, if PLL was bound to only one leaflet, neither D(PLL) nor τ(P) changed. Simple calculations suggest that electrostatic repulsion of the unbound ends prevented neighboring OPg dimers from approaching each other. We believe that a similar mechanism may also operate in cell signaling and that it may e.g. contribute to the controversial results obtained for the ligand driven dimerization of G protein-coupled receptors.


Assuntos
Bicamadas Lipídicas/química , Bicamadas Lipídicas/metabolismo , Peptídeos/química , Peptídeos/metabolismo , Eletricidade Estática , Gramicidina/análogos & derivados , Gramicidina/química , Gramicidina/metabolismo , Ligantes , Polilisina/química , Polilisina/metabolismo , Ligação Proteica
19.
Chem Pharm Bull (Tokyo) ; 59(12): 1481-4, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-22130370

RESUMO

To find candidates with high antimicrobial and low hemolytic activities, many gramicidin S (GS) analogs of various ring sizes have been designed and synthesized. However, syntheses of antimicrobially active analogues of GS having a disordered symmetry structure from C(2) have almost never been reported, because the stable, amphiphilic ß-sheet structure of GS with C(2) symmetry is considered essential for its strong antibacterial activity. In the present studies, novel thirteen cycloundecapeptides 1-13 related to GS were synthesized and examined. Among them, cyclo(-Va1(1)-Orn(2)-Leu(3)-D-Phe(4)-X(5)-Pro(6)-Val(7)-Orn(8)-Leu(9)-D-Phe(10)-Pro(11)-) (X=Lys (10), Orn (11), Arg (12) and Lys(Lys) (13)) resulted in high antibiotic activity against both Gram-positive and Gram-negative microorganisms tested. In addition, 11 showed low toxicity against sheep blood cells compared with that of GS. Further, circular dichroism (CD) spectra of 10-13 had a curve similar to each other, suggesting that the conformations of these analogues in methanol are similar to each other. However, CD spectra of 10-13 were different from that of GS in the 190-210 nm region. These results suggest that the presences of one added amino acid residue at position 5 of 10-13 might be partially effective through a structural change in the biological activity of 10-13. In addition, the structural modifications at position 5 lower the undesirable hemolytic activity and enhance the desirable antibiotic activity.


Assuntos
Antibacterianos/química , Antibacterianos/farmacologia , Bactérias/efeitos dos fármacos , Gramicidina/análogos & derivados , Gramicidina/farmacologia , Hemólise/efeitos dos fármacos , Animais , Antibacterianos/síntese química , Infecções Bacterianas/tratamento farmacológico , Dicroísmo Circular , Gramicidina/síntese química , Humanos , Testes de Sensibilidade Microbiana , Ovinos
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