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1.
J Chem Inf Model ; 63(18): 5823-5833, 2023 09 25.
Artigo em Inglês | MEDLINE | ID: mdl-37684221

RESUMO

Understanding the mechanism of action of the antimicrobial peptide (AMP) in terms of its structure and energetics is the key to designing new potent and selective AMPs. Recently, we reported a membranolytic 14-residue-long lysine-rich cationic antimicrobial peptide (LL-14: NH3+-LKWLKKLLKWLKKL-CONH2) against Pseudomonas aeruginosa, Klebsiella pneumoniae, and Staphylococcus aureus, which is limited by cytotoxicity and expected to undergo facile protease degradation. Aliphatic side-chain-length modification of the cationic amino-acid residues (Lys and Arg) is a popular strategy for designing protease-resistant AMPs. However, the effect of the peptide side-chain length modifications on the membrane binding affinity and its relation to the atomic structure remain an unsolved problem. We report computer simulations that quantitatively calculated the difference in peptide binding affinity to membrane-mimetic-bilayer models (bacterial: 1-palmitoyl-2-oleoyl-phosphatidylethanolamine (POPE)/1-palmitoyl-2-oleoyl-phosphatidylglycerol (POPG) bilayer and mammalian: 1-palmitoyl-2-oleoyl-phosphatidylcholine (POPC) bilayer) upon decreasing or increasing the spacer length of the cationic lysine residues of LL-14 (as well as their arginine analogues). We show that the peptide/bilayer interaction energetics varies drastically in response to spacer length modification. The strength of peptide discrimination depends strongly on the nature of the bilayer (bacterial or mammalian mimetic model). An increase in the lysine spacer length by one carbon (i.e., homolysine analogue of LL-14) is weakly/strongly disfavored by the bacterial/mammalian-membrane-mimetic bilayer. Recently, we have demonstrated an excellent correlation between the antimicrobial activity of the membranolytic cationic peptides and their binding affinity to membrane-mimetic-bilayer models. Thus, the homolysine analogue of LL-14 is a promising noncytotoxic AMP with conserved activity. On the other hand, homoarginine analogue (arginine spacer length increment by a single carbon) was preferred by both the bacteria and the mammalian mimetic bilayers and displayed the strongest affinity for the former among the peptides studied in this work. Thus, the promising most potent homoarginine analogue is likely to be cytotoxic. Shortening the Lys/Arg side chain to a three-carbon spacer (Dab/Agb) improves the binding affinity to bacterial and mammalian-membrane-mimetic bilayers. Arginine and arginine-derivative peptides exhibited stronger binding affinity to the bilayers relative to the lysine analogue. The results provide a plausible explanation to the previous experimental observations, viz., superior antimicrobial activity of the arginine peptides relative to Lys peptides and the improvement of antimicrobial activity upon substitution of Lys with Dab in the cationic peptides. The simulations revealed that the small change in the peptide hydrophobicity by Lys/Arg spacer length modification could drastically alter the energetics of peptide/bilayer binding by fine-tuning the electrostatic interactions. The energetics underlying the peptide selectivity by simple membrane-mimetic bilayer models may be beneficial for designing new selective and protease-resistant AMPs.


Assuntos
Anti-Infecciosos , Lisina , Animais , Homoarginina , Peptídeo Hidrolases , Peptídeos Catiônicos Antimicrobianos/farmacologia , Arginina , Carbono , Cátions , Mamíferos
2.
J Chem Inf Model ; 63(8): 2393-2408, 2023 04 24.
Artigo em Inglês | MEDLINE | ID: mdl-37021489

RESUMO

Understanding the thermodynamics of peptide:membrane binding and the factors that alter the stability is the key to designing potent and selective small antimicrobial peptides. Here, we report the thermodynamics, antimicrobial activity, and mechanism of a de novo designed seven residue long cationic antimicrobial peptide (P4: NH3+-LKWLKKL-CONH2, Charge +4) and its analogs (P5: Lysine's → Arginine's; P6: Lysine's → Uncharged-Histidine's; P7: Tryptophan → Leucine) by combining computation and experiments. Computer simulations predicted the order of decreasing peptide binding affinity to the membrane-mimetic systems (micelle/bilayer) as P5 > P4 > P7 ≫ P6. Antimicrobial assays of these peptides against P. aeruginosa and E. coli at physiological pH 7.4 confirmed P5 as the most potent peptide (followed by P4), whereas P6 showed inferior activity. P7 was found to be inactive against E. coli. Substitution of the uncharged-histidine (P6) by the charged-histidine (P6*) significantly favored micelle/bilayer binding. Thus, P6 was predicted to be an effective antimicrobial peptide only at low pH. Noticeable improvement in the antimicrobial activity of the histidine-peptide (P6) against E. coli (an acid-resistant bacteria) upon lowering the pH was demonstrated and validated the computational claim. The peptides displayed a membranolytic mode of action. The link between the structure and calculated energetics (ΔΔG) has been established, and the correlation between the calculated energetics and the antimicrobial activity has been highlighted. The histidine-peptide (P6) is reported to be active against acid-resistant bacteria, thus, a promising membranolytic pH-sensitive AMP.


Assuntos
Anti-Infecciosos , Histidina , Lisina , Escherichia coli , Micelas , Peptídeos Catiônicos Antimicrobianos/farmacologia , Peptídeos Catiônicos Antimicrobianos/química , Anti-Infecciosos/farmacologia , Anti-Infecciosos/química , Termodinâmica , Concentração de Íons de Hidrogênio , Testes de Sensibilidade Microbiana
3.
Phys Chem Chem Phys ; 24(38): 23669-23678, 2022 Oct 05.
Artigo em Inglês | MEDLINE | ID: mdl-36148810

RESUMO

Antimicrobial peptides (AMPs) are promising antimicrobial and therapeutic agents. Recently, we synthesized a cationic 14 residue AMP (LL-14: LKWLKKLLKWLKKL), which showed high broad-spectrum antimicrobial activity. However, the antimicrobial activity of LL-14 was compromised in the presence of NaCl. Salt sensitivity of antimicrobial potency is one of the fundamental limitations of AMP therapeutics. Thus, understanding the thermodynamics of AMP binding to simple membrane-mimetic systems and the effect of NaCl that contributes to their stability is crucial for designing promising AMPs against microbial infection. In this work, we reported computational analysis of LL-14 binding to SDS micelles (the simplest bacterial membrane mimic) at various NaCl concentrations (0.0%, 0.5%, 1.0% w/v). The thermodynamics of LL-14 dissociation from the SDS micelles was estimated by employing steered molecular dynamics (SMD) simulation followed by umbrella sampling. The results indicated that the increase in NaCl concentration systematically disfavoured the LL-14:SDS binding, primarily by stabilizing the dissociative state (i.e., free LL-14 and free micelles in water). We proposed a kinetic scheme in which the salt-induced selective stabilization of the dissociative state increased the activation barrier for the peptide:micelle binding event resulting in reduced affinity. Center-of-mass pulling indicated that the interactions involving the N-terminal of the LL-14 (residues 1-6) and SDS micelle were crucial for the stability of the LL-14:SDS complex, and LL-14 underwent a conformational change (helix → unstructured) before dissociating from the SDS micelle. The observed structural features from the peptide:micelle dissociation pathway corroborate our previous simulations as well as circular dichroism (CD), and fluorescence experiments.


Assuntos
Anti-Infecciosos , Micelas , Sequência de Aminoácidos , Antibacterianos/farmacologia , Anti-Infecciosos/farmacologia , Peptídeos Catiônicos Antimicrobianos/química , Peptídeos Antimicrobianos , Cloreto de Sódio , Água
4.
J Org Chem ; 86(17): 11310-11323, 2021 09 03.
Artigo em Inglês | MEDLINE | ID: mdl-34479402

RESUMO

The effect of insertion of three geminally dimethyl substituted γ amino acid residues [γ2,2 (4-amino-2,2-dimethylbutanoic acid), γ3,3 (4-amino-3,3-dimethylbutanoic acid), and γ4,4 (4-amino-4,4-dimethylbutanoic acid)] at the (i + 2) position of a two-residue αγ C12 turn segment in a model octapeptide sequence Leu-Phe-Val-Aib-Xxx-Leu-Phe-Val (where Xxx = γ amino acid residues) has been investigated in this study. Solution conformational studies (NMR, CD, and IR) and ab initio calculations indicated that γ3,3 and γ4,4 residues were well accommodated in the ß-hairpin nucleating αγ C12 turns, which gave rise to well-registered hairpins, in contrast to γ2,2, which was unable to form a tight C12 ß-hairpin nucleating turn and promote a well-registered ß-hairpin. Geminal disubstitution at the Cα carbon in γ2,2 led to unfavorable steric contacts, disabling its accommodation in the αγ C12 hairpin nucleating turn unlike the γ3,3 and γ4,4 residues. Geminal substitutions at different carbons along the backbone constrained backbone torsion angles for the three γ amino acid residues differently, generating diverse conformational preferences in them. Folded hairpins were energetically more stable (∼8 to 9 kcal/mol) than the unfolded peptides. Conformational preference of the peptides was independent of the N-terminal protecting group. Such fundamental understanding will instrumentalize the future directed design of foldamers.


Assuntos
Aminoácidos , Peptídeos , Ligação de Hidrogênio , Espectroscopia de Ressonância Magnética , Conformação Proteica , Estrutura Secundária de Proteína
5.
Org Biomol Chem ; 17(11): 3026-3039, 2019 03 13.
Artigo em Inglês | MEDLINE | ID: mdl-30816399

RESUMO

Dicyclohexyl urea (DCU) derivatives of amino acids Fmoc-Phe-DCU (M1), Fmoc-Phg-DCU (M2) and Fmoc-Gaba-DCU (M3) have been shown to form phase selective, thermoreversible and mechanically robust gels in a large range of organic solvents. This is the first report of low molecular weight gelators (LMWG) from DCU derivatives of amino acids. The self-assembly mechanism of the organogels has been probed using concentration dependent 1H NMR, DMSO titration 1H NMR, fluorescence, FTIR, PXRD and FESEM techniques. Self-assembly leading to gelation process is mainly driven by hydrophobicity and π-π stacking interactions in between Fmoc groups. Interestingly, the gels can absorb several kinds of organic dyes efficiently and can be reused for dye absorption for multiple cycles. Additionally, M1-M3 act as sensors for anions like fluoride, acetate and hydroxide, for which they have specific fluorescence response. Gel formation by M1-M3 is completely arrested in the presence of fluoride. The possible binding mode of fluoride has been delineated using DFT studies. Calculations suggest, involvement of urea NH in a six membered intramolecular hydrogen bond, rendering it unavailable for fluoride binding. Backbone -NH of the amino acids of M1-M3 is responsible for fluoride binding. The reported small, economically viable, synthetically facile molecules not only enrich the repertoire of LMWG molecules, but can have multifaceted applications.

6.
Biochim Biophys Acta Biomembr ; 1866(6): 184336, 2024 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-38763273

RESUMO

Short systemic half- life of Antimicrobial Peptides (AMP) is one of the major bottlenecks that limits their successful commercialization as therapeutics. In this work, we have designed analogs of the natural AMP Jelleine, obtained from royal jelly of apis mellifera. Among the designed peptides, J3 and J4 were the most potent with broad spectrum activities against a varied class of ESKAPE pathogens and fungus C. albicans. All the developed peptides were more effective against Gram-negative bacteria in comparison to the Gram-positive pathogens, and were especially effective against P. aeruginosa and C. albicans.J3 and J4 were completely trypsin resistant and serum stable, while retaining the non-cytotoxicity of the parent Jelleine, Jc. The designed peptides were membranolytic in their mode of action. CD and MD simulations in the presence of bilayers, established that J3 and J4 were non-structured even upon membrane binding and suggested that biological properties of the AMPs were innocent of any specific secondary structural requirements. Enhancement of charge to increase the antimicrobial potency, controlling the hydrophobic-hydrophilic balance to maintain non-cytotoxicity and induction of unnatural amino acid residues to impart protease resistance, remains some of the fundamental principles in the design of more effective antimicrobial therapeutics of the future, which may help combat the quickly rising menace of antimicrobial resistance in the microbes.


Assuntos
Peptídeos Catiônicos Antimicrobianos , Candida albicans , Testes de Sensibilidade Microbiana , Peptídeos Catiônicos Antimicrobianos/farmacologia , Peptídeos Catiônicos Antimicrobianos/química , Peptídeos Catiônicos Antimicrobianos/síntese química , Candida albicans/efeitos dos fármacos , Animais , Anti-Infecciosos/farmacologia , Anti-Infecciosos/química , Bactérias Gram-Negativas/efeitos dos fármacos , Peptídeo Hidrolases/química , Peptídeo Hidrolases/metabolismo , Humanos , Abelhas , Peptídeos Antimicrobianos/farmacologia , Peptídeos Antimicrobianos/química , Simulação de Dinâmica Molecular , Oligopeptídeos
7.
ACS Infect Dis ; 10(2): 562-581, 2024 02 09.
Artigo em Inglês | MEDLINE | ID: mdl-38294842

RESUMO

Antimicrobial peptides (AMPs) have been an alternate promising class of therapeutics in combating global antibiotic resistance threat. However, the short half-life of AMPs, owing to protease degradability, is one of the major bottlenecks in its commercial success. In this study, we have developed all-D-amino acid containing small cationic peptides P4C and P5C, which are completely protease-resistant, noncytotoxic, nonhemolytic, and potent against the ESKAPE pathogens in comparison to their L analogues. MD simulations suggested marginal improvement in the peptide-binding affinity to the membrane-mimetic SDS micelle (∼ 1 kcal/mol) in response to L → D conversion, corroborating the marginal improvement in the antimicrobial activity. However, L → D chirality conversion severely compromised the peptide:protease (trypsin) binding affinity (≥10 kcal/mol). The relative distance between the scissile peptide carbonyl and the catalytic triad of the protease (H57, D102, and S195) was found to be significantly altered in the D-peptide:protease complex (inactive conformation) relative to the active L-peptide:protease complex. Thus, the poor binding affinity between D-peptides and the protease, resulting in the inactive complex formation, explained their experimentally observed proteolytic stability. This mechanistic insight might be extended to the proteolytic stability of the D-peptides in general and stimulate the rational design of protease-resistant AMPs.


Assuntos
Anti-Infecciosos , Peptídeo Hidrolases , Peptídeo Hidrolases/metabolismo , Aminoácidos , Peptídeos Catiônicos Antimicrobianos/farmacologia , Peptídeos Catiônicos Antimicrobianos/química , Anti-Infecciosos/química , Endopeptidases
8.
ACS Omega ; 8(39): 36370-36385, 2023 Oct 03.
Artigo em Inglês | MEDLINE | ID: mdl-37810672

RESUMO

Chirality is an omnipresent feature in nature's architecture starting from simple molecules like amino acids to complex higher-order structures viz. proteins, DNA, and RNA. The L configuration of proteinogenic amino acids gives rise to right-handed helices. Ambidexterity is as rare in organisms as in molecules. There are only a few reports of ambidexterity in single-peptide molecules composed of either mixed L and D or achiral residues. Here, we report, for the first time, the ambidextrous and left-handed helical conformations in the chiral nonapeptides P1-P3 (Boc-LUVUγx,xULUV-OMe where U = Aib, x,x = 2,2/3,3/4,4), containing chiral L α amino acid residues, in addition to the usually observed right-handed helical conformation. The centrally located achiral γ residue, capable of adopting both left and right-handed helical conformations, induces its handedness on the neighboring chiral and achiral residues, leading to the observation of both left and right-handed helices in P2 and P3. The presence of a single water molecule proximal to the γ residue induces the reversal of helix handedness by forming distinct and stable water-mediated hydrogen bonds. This gives rise to ambidextrous helices as major conformers in P1 and P2. The absence of the observation of ambidexterity in P3 might be due to the inability of γ4,4 in the recruitment of a water molecule. Experiments (NMR, X-ray, and CD) and density functional theory (DFT) calculations suggest that the position of geminal disubstitution is crucial for determining the population of the amenable helical conformations (ambidextrous, left and right-handed) in these chiral peptides.

9.
J Phys Chem B ; 126(28): 5262-5273, 2022 07 21.
Artigo em Inglês | MEDLINE | ID: mdl-35815580

RESUMO

Recently, we had reported a synthetic positively charged leucine-rich 14-residue-long antimicrobial peptide (AMP, LL-14: NH3+-LKWLKKLLKWLKKL-CONH2), which was highly active and cytotoxic relative to its valine analogue (VV-14). However, the thermodynamics underlying this differential toxicity and antimicrobial activity was unclear. Understanding the energetics of peptide binding to micelles (simplest membrane mimic, viz., SDS as a bacterial membrane and DPC as a eukaryotic membrane) and the effect of Leu → Val peptide mutations on the stability of the peptide:micelle complexes are of great academic interest and relevant for the rational design of potent and selective AMPs for therapeutic use. Here, we have reported the molecular dynamics free energy simulations that allowed us to quantitatively estimate the strength of peptide discrimination (based on single- or multiple-site Leu/Val mutations in LL-14) by membrane mimetic micelles (SDS and DPC) and decipher the energetics underlying peptide selectivity by micelles. The Leu-containing peptide (LL-14) was found to be preferred for micelle (SDS and DPC) binding relative to its Val analogues (single or multiple Val mutants). The strength of the preference depended on the position of the Leu/Val mutation in the peptide. Surprisingly, the N-terminal LL-14 single mutation (Leu → Val: L1V) was found to fine-tune the electrostatic interactions, resulting in the highest peptide selectivity (ΔΔG ∼ 8 kcal/mol for both SDS and DPC). However, the mechanism of L1V peptide selectivity was distinctly different for SDS and DPC micelles. SDS ensured high selectivity by disrupting the peptide:micelle salt bridge, whereas DPC desolvated the broken-peptide-backbone hydrogen bond in the V1 peptide:micelle complex. Mutations (Leu → Val) in the middle positions of the LL-14 (4th, 7th, 8th, and 11th) were disfavored by the micelles primarily due to the loss of peptide:micelle hydrophobic interactions. Peptides differing at the C-terminal (i.e., L14V) were recognized by SDS micelles (ΔΔG ∼ 4 kcal/mol) by altering peptide:micelle interactions. L14V mutation, on the other hand, did not play any role in the peptide:DPC binding, as no direct interactions between the C-terminal and DPC micelle were observed due to obvious electrostatic reasons. The strength of selectivity favoring LL-14 binding against VV-14 was found to be much higher for DPC micelles (ΔΔG ∼ 25 kcal/mol) relative to SDS micelles (ΔΔG ∼ 19 kcal/mol). The loss of the peptide:micelle hydrophobic contact in response to LL-14 → VV-14 mutation was found to be significantly larger for DPC relative to SDS micelles, resulting in higher discriminatory power for the former. Peptide:SDS salt bridges seemed to prevent the loss of peptide:micelle hydrophobic contact to some extent, leading to weaker selectivity for SDS micelles. High selectivity of DPC micelles provided an efficient mechanism for VV-14 dissociation from DPC micelles, whereas low-selectivity of SDS micelles ensured binding of both LL-14 and VV-14. To the best of our knowledge, this is the first study in which the experimental observations (antimicrobial activity and toxicity) between leucine-rich and valine-rich peptides have been explained by establishing a direct link between the energetics and structures.


Assuntos
Anti-Infecciosos , Micelas , Anti-Infecciosos/química , Anti-Infecciosos/farmacologia , Peptídeos Catiônicos Antimicrobianos/química , Peptídeos Catiônicos Antimicrobianos/genética , Peptídeos Catiônicos Antimicrobianos/farmacologia , Peptídeos Antimicrobianos , Leucina/genética , Mutação , Ressonância Magnética Nuclear Biomolecular , Fosforilcolina/química , Dodecilsulfato de Sódio/química , Valina
10.
ACS Omega ; 7(18): 15951-15968, 2022 May 10.
Artigo em Inglês | MEDLINE | ID: mdl-35571791

RESUMO

Rapidly growing antimicrobial resistance (AMR) against antibiotics has propelled the development of synthetic antimicrobial peptides (AMPs) as potential antimicrobial agents. An antimicrobial peptide Nle-Dab-Trp-Nle-Dab-Dab-Nle-CONH2 (P36; Nle = norleucine, Dab = diaminobutyric acid, Trp = tryptophan) potent against Pseudomonas aeruginosa (P. aeruginosa) has been developed in the present study. Rational design strategy adopted in this study led to the improvisation of the therapeutic qualities such as activity, salt tolerance, cytotoxicity, and protease resistance of the template peptide P4, which was earlier reported from our group. P36 exhibited salt tolerant antimicrobial potency against P. aeruginosa, along with very low cytotoxicity against mammalian cell lines. P36 was found to be nonhemolytic and resistant toward protease degradation which qualified it as a potent antimicrobial agent. We have investigated the mechanism of action of this molecule in detail using several experimental techniques (spectroscopic, biophysical, and microscopic) and molecular dynamics simulations. P36 was a membrane active AMP with membrane destabilization and deformation abilities, leading to leakage of the intracellular materials and causing eventual cell death. The interaction between P36 and the microbial membrane/membrane mimics was primarily driven by electrostatics. P36 was unstructured in water and upon binding to the microbial membrane mimic SDS, suggesting no influence of secondary structure on its antimicrobial potency. Positive charge, optimum hydrophobic-hydrophilic balance, and chain length remained the most important concerns to be addressed while designing small cationic antimicrobial peptides.

11.
Chem Asian J ; 17(14): e202200356, 2022 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-35603989

RESUMO

We studied the effect of variable backbone dimethyl-substitution of γ amino acid residues (γ2,2 , γ3,3 and γ4,4 ) on the conformation and assembly, in crystals and solution of their Fmoc derivatives. The crystal structure of γ2,2 and γ4,4 derivatives showed distinct conformations (open/close for γ2,2 /γ4,4 ) that differed in torsion angles, hydrogen-bonding and most importantly the π-π Fmoc-stacking interactions (relatively favourable for γ4,4 -close). Fmoc derivatives existed in an equilibrium between major-monomeric (low energy, non-hydrogen bonded) and minor-dimeric (high energy, hydrogen bonded) populations in solution. The rate of major/minor population exchange was dependent on the position of substitution, highest being for γ4,4 derivative. In solution, assembly of Fmoc derivatives was solvent dependent, but it was independent of the position of geminal substitution. Crystallization was primarily governed by the stabilization of high-energy dimer by favourable π-π stacking involving Fmoc moieties. High free-energy of the dimers (γ2,2 -close, γ3,3 -open/close) offset favourable stacking interactions and hindered crystallization.


Assuntos
Aminoácidos , Fluorenos , Aminoácidos/química , Fluorenos/química , Ligação de Hidrogênio , Conformação Molecular , Solventes
12.
Front Chem ; 9: 691532, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34222199

RESUMO

Rapid rise of antimicrobial resistance against conventional antimicrobials, resurgence of multidrug resistant microbes and the slowdown in the development of new classes of antimicrobials, necessitates the urgent development of alternate classes of therapeutic molecules. Antimicrobial peptides (AMPs) are small proteins present in different lifeforms in nature that provide defense against microbial infections. They have been effective components of the host defense system for a very long time. The fact that the development of resistance by the microbes against the AMPs is relatively slower or delayed compared to that against the conventional antibiotics, makes them prospective alternative therapeutics of the future. Several thousands of AMPs have been isolated from various natural sources like microorganisms, plants, insects, crustaceans, animals, humans, etc. to date. However, only a few of them have been translated commercially to the market so far. This is because of some inherent drawbacks of the naturally obtained AMPs like 1) short half-life owing to the susceptibility to protease degradation, 2) inactivity at physiological salt concentrations, 3) cytotoxicity to host cells, 4) lack of appropriate strategies for sustained and targeted delivery of the AMPs. This has led to a surge of interest in the development of synthetic AMPs which would retain or improve the antimicrobial potency along with circumventing the disadvantages of the natural analogs. The development of synthetic AMPs is inspired by natural designs and sequences and strengthened by the fusion with various synthetic elements. Generation of the synthetic designs are based on various strategies like sequence truncation, mutation, cyclization and introduction of unnatural amino acids and synthons. In this review, we have described some of the AMPs isolated from the vast repertoire of natural sources, and subsequently described the various synthetic designs that have been developed based on the templates of natural AMPs or from de novo design to make commercially viable therapeutics of the future. This review entails the journey of the AMPs from their natural sources to the laboratory.

13.
RSC Adv ; 11(58): 36836-36849, 2021 Nov 10.
Artigo em Inglês | MEDLINE | ID: mdl-35494385

RESUMO

Recently, we reported a cationic 14 residue peptide LL-14 (LKWLKKLLKWLKKL) with salt-sensitive broad-spectrum antimicrobial potency. However, the mechanism of its salt (NaCl) sensitivity remained unclear. In this study, we have reported computational (∼14.2 µs of MD) and experimental (CD, fluorescence) investigations to examine the salt-sensitivity and the role of peptide secondary structure on LL-14 binding to simple membrane mimetic (SDS, DPC) systems. LL-14 was shown to adopt a random coil (Pc) conformation in water and α-helical conformation (Ph) in the peptide:SDS micelle complex, accompanied by tryptophan burial, using both simulations and experiments. Simulations successfully deconvoluted the LL-14:micelle binding event in terms of secondary structure (random coil Pc versus helix Ph) and gave atomic insight into the initial and final LL-14:SDS complexes. Electrostatics drove the N-terminus (L1 and K2) of LL-14 (Pc or Ph) to bind the SDS micellar surface, initiating complex formation. LL-14 in amphipathic Ph conformation bound faster and buried deeper into the SDS micelle relative to Pc. Increasing NaCl concentration incrementally delayed LL-14:micelle binding by shielding the overall charges of the interacting partners. LL-14 binding to the SDS micelle was significantly faster relative to that of the zwitterionic DPC micelle due to electrostatic reasons. Cationic α-helical amphipathic peptides (with positively charged N-terminus) with low salt-ion concentration seemed to be ideal for faster SDS binding.

14.
ChemMedChem ; 16(2): 355-367, 2021 01 19.
Artigo em Inglês | MEDLINE | ID: mdl-33026188

RESUMO

Herein we report the efficacy and toxicity of three de novo designed cationic antimicrobial peptides (AMPs) LL-14, VV-14 and ßß-14, where side chains of the hydrophobic amino acids were reduced gradually. The AMPs showed broad-spectrum antimicrobial activity against three pathogens from the ESKAPE group and two fungal strains. This study showed that side chains which are either too long or too short increase toxicity and lower antimicrobial activity, respectively. VV-14 was found to be non-cytotoxic and highly potent under physiological salt concentrations against several pathogens, especially Salmonella typhi TY2. These AMPs acted via membrane deformation, depolarization, and lysis. The activity of the AMPs is related to their ability to take on amphipathic helical conformations in the presence of microbial membrane mimics. Among AMPs with the same charge, hydrophobic interactions between the side chains of the residues with cell membrane lipids determine their antimicrobial potency and cytotoxicity. Strikingly, an optimum hydrophobic interaction is the crux of generating highly potent non-cytotoxic AMPs.


Assuntos
Aminoácidos/farmacologia , Antibacterianos/farmacologia , Antifúngicos/farmacologia , Peptídeos Catiônicos Antimicrobianos/farmacologia , Bactérias/efeitos dos fármacos , Fungos/efeitos dos fármacos , Aminoácidos/química , Antibacterianos/química , Antifúngicos/química , Peptídeos Catiônicos Antimicrobianos/química , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Humanos , Interações Hidrofóbicas e Hidrofílicas , Testes de Sensibilidade Microbiana , Estrutura Secundária de Proteína
15.
Acc Chem Res ; 42(10): 1628-39, 2009 Oct 20.
Artigo em Inglês | MEDLINE | ID: mdl-19572698

RESUMO

Nature has used the all-alpha-polypeptide backbone of proteins to create a remarkable diversity of folded structures. Sequential patterns of 20 distinct amino acids, which differ only in their side chains, determine the shape and form of proteins. Our understanding of these specific secondary structures is over half a century old and is based primarily on the fundamental elements: the Pauling alpha-helix and beta-sheet. Researchers can also generate structural diversity through the synthesis of polypeptide chains containing homologated (omega) amino acid residues, which contain a variable number of backbone atoms. However, incorporating amino acids with more atoms within the backbone introduces additional torsional freedom into the structure, which can complicate the structural analysis. Fortunately, gabapentin (Gpn), a readily available bulk drug, is an achiral beta,beta-disubstituted gamma amino acid residue that contains a cyclohexyl ring at the C(beta) carbon atom, which dramatically limits the range of torsion angles that can be obtained about the flanking C-C bonds. Limiting conformational flexibility also has the desirable effect of increasing peptide crystallinity, which permits unambiguous structural characterization by X-ray diffraction methods. This Account describes studies carried out in our laboratory that establish Gpn as a valuable residue in the design of specifically folded hybrid peptide structures. The insertion of additional atoms into polypeptide backbones facilitates the formation of intramolecular hydrogen bonds whose directionality is opposite to that observed in canonical alpha-peptide helices. If hybrid structures mimic proteins and biologically active peptides, the proteolytic stability conferred by unusual backbones can be a major advantage in the area of medicinal chemistry. We have demonstrated a variety of internally hydrogen-bonded structures in the solid state for Gpn-containing peptides, including the characterization of the C(7) and C(9) hydrogen bonds, which can lead to ribbons in homo-oligomeric sequences. In hybrid alphagamma sequences, distinct C(12) hydrogen-bonded turn structures support formation of peptide helices and hairpins in longer sequences. Some peptides that include the Gpn residue have hydrogen-bond directionality that matches alpha-peptide helices, while others have the opposite directionality. We expect that expansion of the polypeptide backbone will lead to new classes of foldamer structures, which are thus far unknown to the world of alpha-polypeptides. The diversity of internally hydrogen-bonded structures observed in hybrid sequences containing Gpn shows promise for the rational design of novel peptide structures incorporating hybrid backbones.


Assuntos
Aminas/química , Ácidos Cicloexanocarboxílicos/química , Desenho de Fármacos , Peptídeos/química , Ácido gama-Aminobutírico/química , Gabapentina , Ligação de Hidrogênio , Conformação Proteica , Estereoisomerismo
16.
RSC Adv ; 10(9): 5220-5233, 2020 Jan 29.
Artigo em Inglês | MEDLINE | ID: mdl-35498311

RESUMO

The self-assembly of a series of low molecular weight gelator dipeptides containing para amino benzoic acid has been studied in mechanistic detail. All four dipeptides form phase selective, thermoreversible, rigid gels in a large range of organic solvents and fuels such as petrol, diesel, and kerosene. The mechanism of self-assembly has been dissected in detail using several experimental techniques. Self-assembly is driven mainly by aromatic and hydrophobic interactions. Hydrogen bonding groups, though present, seem to make a trivial contribution towards the self-assembly process. Phase selective gelation abilities in fuels in the presence of acidic, basic and saline conditions, together with the easy recovery of fuels from the organogels, render the peptides potential candidates for addressing oil-spill recovery. Being electron rich systems, these organogelators can absorb cationic dyes with >90% efficiency from wastewater. Finally, conducting biomaterials have been synthesized by the insertion of reduced graphene oxide into the organogels. Such small peptide based gelator molecules, being economically viable and easy to prepare, in addition to being multifunctional, are a hot area of research in the field of materials chemistry.

17.
RSC Adv ; 10(69): 42062-42075, 2020 Nov 17.
Artigo em Inglês | MEDLINE | ID: mdl-35516776

RESUMO

Learning from nature, molecular self-assembly has been used extensively to generate interesting materials using a bottom up approach. The enthusiasm in this field of research stems from the unique properties of these materials and their diverse applications. The field has not been limited to studying assembly of similar types of molecules but extended to multi component systems via the co-assembly phenomenon. We have designed two charge complementary peptides to study their co-assembly in mechanistic detail in the present work. The cooperative self-assembly is mainly driven by electrostatic interaction that is aided by aromatic interactions, hydrogen bonding interactions and hydrophobic interactions. The hydrogels obtained have been employed in waste water remediation. Both the self-assembled and co-assembled hydrogels are capable of removal of different kinds of organic dyes (cationic, anionic and neutral) and toxic metal ions (Ni2+, Co2+, Pb2+ and Hg2+) individually and as a mixture from water with high efficiency. Additionally, the peptides developed in this study can act as ion sensors and detect arsenic in its most toxic (III/V) oxidation states. Molecular understanding of the assembly process is of fundamental importance in the rational design of such simple, robust yet economically viable materials with versatile and novel applications.

18.
ACS Appl Bio Mater ; 3(9): 6251-6262, 2020 Sep 21.
Artigo em Inglês | MEDLINE | ID: mdl-35021757

RESUMO

A charged synthetic peptide-based noncytotoxic hydrogelator was employed in encapsulation, storage, and sustainable release of different kinds of drugs, namely, ciprofloxacin (CP), an antibiotic; 5-fluorouracil (5-FU), an anticancer drug and proteins like lysozyme and bovine serum albumin (BSA). Hydrogelation of the peptide and its coassembly with the drug molecules were studied to obtain mechanistic details. All of the different cargos were capable of sustained and efficient release from the delivery platform. The drugs were found to retain their activity post release, while the proteins showed complete retention of their secondary structure. While about 80% CP was released at physiological pH over a period of 3 days, 5-FU was better released (73%) at an acidic pH (5.5) in comparison to the physiological pH (68%). Lysozyme was better released (82%) than BSA (43%) owing to the smaller size of the former and negative charge on the latter. Such biocompatible multicargo-releasing platforms from simple economically viable biomaterials, capable of sustained and tissue-specific release of cargo, are extremely promising in topical delivery of therapeutics.

19.
Biochim Biophys Acta Biomembr ; 1862(4): 183177, 2020 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-31954105

RESUMO

The widespread abuse of antibiotics has led to the use of antimicrobial peptides (AMPs) as a replacement for the existing conventional therapeutic agents for combating microbial infections. The broad-spectrum activity and the resilient nature of AMPs has mainly aggrandized their utilization. Here, we report the design of non-toxic, non-hemolytic and salt tolerant undecapeptides (AMP21-24), derived by modification of a peptide P5 (NH2-LRWLRRLCONH2) reported earlier by our group. Our results depict that the designed peptides show potency against several bacterial as well as fungal strains. Circular dichroism (CD) spectroscopy in combination with molecular dynamic (MD) simulations confirm that the peptides are unstructured. Intrinsic tryptophan fluorescence quenching as well as interaction studies using isothermal calorimetry (ITC) of these peptides in the presence of biological microbial membrane mimics establish the strong microbial membrane affinity of these AMPs. Membrane permeabilization assay and cytoplasmic membrane depolarization studies of Pseudomonas aeruginosa and Candida albicans in the presence of AMPs also hint towards the AMP-membrane interactions. Leakage of calcein dye from membrane mimic liposomes, live cell NMR and field emission scanning electron microscopy (FESEM) studies suggest that the AMPs may be primarily involved in membrane perturbation leading to release of intracellular substances resulting in subsequent microbial cell death. Confocal laser scanning microscopy (CLSM) shows localization of the peptides throughout the cell, indicating the possibility of secondary mode of actions. Electrostatic interactions seem to govern the preferential binding of the AMPs to the microbial membranes in comparison to the mammalian membranes as seen from the MD simulations.


Assuntos
Peptídeos Catiônicos Antimicrobianos/farmacologia , Infecções Bacterianas/tratamento farmacológico , Permeabilidade da Membrana Celular/efeitos dos fármacos , Membrana Celular/efeitos dos fármacos , Sequência de Aminoácidos/genética , Peptídeos Catiônicos Antimicrobianos/química , Infecções Bacterianas/microbiologia , Calorimetria , Candida albicans/efeitos dos fármacos , Candida albicans/patogenicidade , Membrana Celular/química , Membrana Celular/ultraestrutura , Dicroísmo Circular , Humanos , Testes de Sensibilidade Microbiana , Microscopia Confocal , Microscopia Eletroquímica de Varredura , Simulação de Dinâmica Molecular , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/patogenicidade , Eletricidade Estática , Relação Estrutura-Atividade
20.
J Am Chem Soc ; 131(16): 5956-65, 2009 Apr 29.
Artigo em Inglês | MEDLINE | ID: mdl-19341285

RESUMO

Hybrid peptide segments containing contiguous alpha and gamma amino acid residues can form C(12) hydrogen bonded turns which may be considered as backbone expanded analogues of C(10) (beta-turns) found in alphaalpha segments. Exploration of the regular hydrogen bonded conformations accessible for hybrid alphagamma sequences is facilitated by the use of a stereochemically constrained gamma amino acid residue gabapentin (1-aminomethylcyclohexaneacetic acid, Gpn), in which the two torsion angles about C(gamma)-C(beta) (theta(1)) and C(beta)-C(alpha) (theta(2)) are predominantly restricted to gauche conformations. The crystal structures of the octapeptides Boc-Gpn-Aib-Gpn-Aib-Gpn-Aib-Gpn-Aib-OMe (1) and Boc-Leu-Phe-Val-Aib-Gpn-Leu-Phe-Val-OMe (2) reveal two distinct conformations for the Aib-Gpn segment. Peptide 1 forms a continuous helix over the Aib(2)-Aib(6) segment, while the peptide 2 forms a beta-hairpin structure stabilized by four cross-strand hydrogen bonds with the Aib-Gpn segment forming a nonhelical C(12) turn. The robustness of the helix in peptide 1 in solution is demonstrated by NMR methods. Peptide 2 is conformationally fragile in solution with evidence of beta-hairpin conformations being obtained in methanol. Theoretical calculations permit delineation of the various C(12) hydrogen bonded structures which are energetically feasible in alphagamma and gammaalpha sequences.


Assuntos
Aminas/química , Ácidos Cicloexanocarboxílicos/química , Peptídeos/química , Ácido gama-Aminobutírico/química , Simulação por Computador , Cristalografia por Raios X , Gabapentina , Ligação de Hidrogênio , Modelos Moleculares , Peptídeos/síntese química , Conformação Proteica , Estrutura Secundária de Proteína , Estereoisomerismo
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