RESUMEN
Antimicrobial peptides (AMPs) constitute a promising tool in the development of novel therapeutic agents useful in a wide range of bacterial and fungal infections. Among the modifications improving pharmacokinetic and pharmacodynamic characteristics of natural AMPs, an important role is played by lipidation. This study focuses on the newly designed and synthesized lipopeptides containing multiple Lys residues or their shorter homologues with palmitic acid (C16) attached to the side chain of a residue located in the center of the peptide sequence. The approach resulted in the development of lipopeptides representing a model of surfactants with two polar headgroups. The aim of this study is to explain how variations in the length of the peptide chain or the hydrocarbon side chain of an amino acid residue modified with C16, affect biological functions of lipopeptides, their self-assembling propensity, and their mode of action.
Asunto(s)
Péptidos Catiónicos Antimicrobianos/química , Infecciones Bacterianas/tratamiento farmacológico , Lipopéptidos/química , Micosis/tratamiento farmacológico , Secuencia de Aminoácidos/genética , Péptidos Catiónicos Antimicrobianos/genética , Péptidos Catiónicos Antimicrobianos/farmacología , Infecciones Bacterianas/microbiología , Candida albicans/efectos de los fármacos , Candida albicans/patogenicidad , Escherichia coli/efectos de los fármacos , Humanos , Lipopéptidos/genética , Lipopéptidos/farmacología , Pruebas de Sensibilidad Microbiana , Micosis/microbiología , Relación Estructura-ActividadRESUMEN
Short cationic antimicrobial lipopeptides with surfactant-like structure are promising antibiotic candidates that preferentially target microbial membranes. Therefore, we focused our study on double-chain lipopeptides, (C10-16)2Dab-KKK-NH2 and (C10-16)2Dap-KKK-NH2, where Dab and Dap are 2,4-diaminobutyric and 2,3-diaminopropionic acids, respectively. We tried to answer a question how the self-assembly behaviour affects biological activities of the tested compounds. The subject compounds were synthesized by solid-phase method and screened for their antimicrobial and haemolytic activities. Cytotoxicity tests on human keratinocytes were carried out for the most promising lipopeptides. Self-assembly properties were evaluated by both experimental and theoretical methods. Interactions with membrane models were examined using the ITC and FTIR techniques. All the lipopeptides studied showed the tendency to self-assembly in solution, and this behaviour was affected by the length of the hydrocarbon chains. Acyl chain elongation supported the formation of the bilayer structure and deprived the lipopeptides of antimicrobial activity. A multi-step mechanism of interaction with a negatively charged membrane was observed for the short-chain lipopeptides, indicating other processes accompanying the binding process. Short-chain lipopeptides were able to penetrate into the liposome's interior and/or cause the rupture of the liposome, this being compatible with their high antimicrobial activity.
Asunto(s)
Antibacterianos/química , Antibacterianos/farmacología , Lipopéptidos/química , Lipopéptidos/farmacología , Péptidos Catiónicos Antimicrobianos/química , Péptidos Catiónicos Antimicrobianos/farmacología , Bacterias/efectos de los fármacos , Infecciones Bacterianas/tratamiento farmacológico , Candida albicans/efectos de los fármacos , Candidiasis/tratamiento farmacológico , Hemólisis/efectos de los fármacos , Humanos , Membrana Dobles de Lípidos/metabolismo , Liposomas/metabolismo , Pruebas de Sensibilidad Microbiana , Simulación de Dinámica MolecularRESUMEN
This study investigates short cationic antimicrobial lipopeptides composed of 2-4 amino acid residues and C12-C18 fatty acids attached to the N-terminal part of the peptides. The findings were discussed in the context of the relationship among biological activity, self-assembly, stability, and membrane interactions. All the lipopeptides showed the ability to self-assemble in PBS solution. In most cases, the critical aggregation concentration (CAC) much surpassed the minimal inhibitory concentration (MIC) values, suggesting that monomers are the main active form of lipopeptides. The introduction of ß-alanine into the peptide sequence resulted in a compound with a high propensity to fibrillate, which increased the peptide stability and activity against S. epidermidis and C. albicans and reduced the cytotoxicity against human keratinocytes. The results of our study indicated that the target of action of lipopeptides is the bacterial membrane. Interestingly, the type of peptide counterion may affect the degree of penetration of the lipid bilayer. In addition, the binding of the lipopeptide to the membrane of Gram-negative bacteria may lead to the release of calcium ions necessary for stabilization of the lipopolysaccharide layer.
RESUMEN
A systematic DFT conformational studies of four building blocks of TNA with cytosine attached to the C1' atom of the α-L-threofuranose moiety are presented. Structures bearing 2'-OR and 3'-OR substituents, where R represents H, CH3 and phosphate groups, were used in the studies using a B3LYP functional in the gas phase. The χ angle (C2-N1-C1'-O4'), the ν0-ν4 endocyclic torsion angles and the exocyclic torsion angles ε (X-O2'-C2'-C1') and γ (X-O3'-C3'-C2') geometry parameter variations were taken into consideration. Three energy minima, high-anti, anti and syn, were found for the rotation about the C1'-N1 bond. The high-anti orientation of the base with respect to the sugar moiety, turned out to be preferred, regardless of the substituents at the C2' and C3' positions. Other orientations are at least 1.65â¯kcal/mol higher in Gibbs free energy than the high-anti one. It has been shown that intramolecular H-bonds and the anomeric effect of phosphate groups strongly affect the conformational preferences of the studied compounds. Further, the structure of substituents attached to the sugar moiety influence the pucker of the furanoid ring. The furanoid ring in the global minima of the compound with two OH groups (TC1) in the 2' and 3' positions, and the compound having a 3'-phosphate group (TC2), adopt roughly the same conformation located at the southern range of the pseudorotation wheel, and thus are close to those found in the B type DNA helix. The low-energy high-anti rotamers of the geometry with the phosphate group attached to the sugar ring in the 2' position (TC3) and the geometry with two methoxyl groups (TC4) have their furanoid rings in conformations resembling those found in A DNA and RNA helices (the northern range of the pseudorotation wheel).
Asunto(s)
Citidina/química , Modelos Moleculares , Conformación de Ácido Nucleico , Ácidos Nucleicos/química , Citidina/análogos & derivados , Enlace de Hidrógeno , Estructura Molecular , Relación Estructura-ActividadRESUMEN
In this paper, we examine antimicrobial and cytotoxic activities, self-assembly and interactions with anionic and zwitterionic membranes of short arginine-rich lipopeptides: C16-RRRR-NH2, C14-RRRR-NH2, C12-RRRR-NH2, and C16-PRRR-NH2. They show a tendency to self-assembly into micelles, but it is not required for antimicrobial activity. The membrane binding of the lipopeptides can be accompanied by other factors such as: peptide aggregation, pore formation or micellization of phospholipid bilayer. The shortening of the acyl chain results in compounds with a lower haemolytic activity and a slightly improved antimicrobial activity against Gram-positive bacteria, what indicates enhanced cell specificity. Results of coarse-grained molecular dynamics simulations indicate different organization of membrane lipids upon binding of arginine-based lipopeptides and the previously studied lysine-based ones.