Combinatorial Synthesis and Directed Evolution Applied to the Production of &amp;#945;-Helix Forming Antimicrobial Peptides Analogues

Castro, Mariana S.; Cilli, Eduardo Maffud; Fontes, Wagner

doi:10.2174/138920306779025648

Cited by 28 publications

(19 citation statements)

References 40 publications

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“…These results suggested that A-hBD-2 is more effective for wound healing at the same peptide concentration under the same wound conditions. It is worth noting that the N-terminal peptide was modified and an α-helix structure added, and this may have been important in improving the stability of peptides [17]. Changing the amino acid sequence on the N-and C-terminals of LL-37 decreased its cytotoxicity and enhance its structural stability and antibacterial activity [18].…”

Section: Discussionmentioning

confidence: 99%

The Designer Antimicrobial Peptide A-hBD-2 Facilitates Skin Wound Healing by Stimulating Keratinocyte Migration and Proliferation

Mi¹,

Liu²,

Liu³

et al. 2018

Cell Physiol Biochem

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Background/Aims: Antimicrobial peptides are effective promoters of wound healing but are susceptible to degradation. In this study, we replaced the GIGDP unit on the N-terminal of the endogenous human antimicrobial peptide hBD-2 with APKAM to produce A-hBD-2 and analyzed the effect on wound healing both in vitro and in vivo. Methods: The effects of A-hBD-2 and hBD-2 on cytotoxicity and proliferation in keratinocytes were assessed by Cell Counting Kit-8 assay. The structural stability and antimicrobial activity of hBD-2 and A-hBD-2 were evaluated against Staphylococcus aureus. RNA and proteins levels were evaluated by real-time PCR and western blotting, respectively. Cell migration was evaluated using a transwell assay. Cell cycle analysis was performed by flow cytometry. Wound healing was assessed in Sprague-Dawley rats. Epidermal thickness was evaluated by hematoxylin and eosin staining. Results: We found that hBD-2 exhibited cytotoxicity at high concentrations and decreased the structural stability in the presence of high sodium chloride concentrations. A-hBD-2 exhibited increased structural stability and antimicrobial activity, and had lower cytotoxicity in keratinocytes. A-hBD-2 increased the migration and proliferation of keratinocytes via phosphorylation of EGFR and STAT3 and suppressed terminal differentiation of keratinocytes. We also found that A-hBD-2 elicited mobilization of intracellular Ca²⁺ and stimulated keratinocytes to produce pro- and anti-inflammatory cytokines and chemokines via phospholipase C activation. Furthermore, A-hBD-2 promoted wound healing in vivo. Conclusion: Our data suggest that A-hBD-2 may be a promising candidate therapy for wound healing.

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Section: Discussionmentioning

confidence: 99%

The Designer Antimicrobial Peptide A-hBD-2 Facilitates Skin Wound Healing by Stimulating Keratinocyte Migration and Proliferation

Mi¹,

Liu²,

Liu³

et al. 2018

Cell Physiol Biochem

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“…cAMPs are characterized by the high occurrence of basic amino acids, a considerable percentage of hydrophobic amino acids and a greater tendency to adopt amphipathic α-helical structures. Unlike classical antibiotics, which attack specific enzymes or receptors in the cell [4], cAMPs change membrane permeability and promote perturbation in the pathogen cell membrane, which is the basis of their mechanism of action [5].…”

Section: Introductionmentioning

confidence: 99%

Dynamics and Conformational Studies of TOAC Spin Labeled Analogues of Ctx(Ile21)-Ha Peptide from Hypsiboas albopunctatus

et al. 2013

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Antimicrobial peptides (AMPs) isolated from several organisms have been receiving much attention due to some specific features that allow them to interact with, bind to, and disrupt cell membranes. The aim of this paper was to study the interactions between a membrane mimetic and the cationic AMP Ctx(Ile21)-Ha as well as analogues containing the paramagnetic amino acid 2,2,6,6-tetramethylpiperidine-1-oxyl-4-amino-4-carboxylic acid (TOAC) incorporated at residue positions n = 0, 2, and 13. Circular dichroism studies showed that the peptides, except for [TOAC13]Ctx(Ile21)-Ha, are unstructured in aqueous solution but acquire different amounts of α-helical secondary structure in the presence of trifluorethanol and lysophosphocholine micelles. Fluorescence experiments indicated that all peptides were able to interact with LPC micelles. In addition, Ctx(Ile21)-Ha and [TOAC13]Ctx(Ile21)-Ha peptides presented similar water accessibility for the Trp residue located near the N-terminal sequence. Electron spin resonance experiments showed two spectral components for [TOAC0]Ctx(Ile21)-Ha, which are most likely due to two membrane-bound peptide conformations. In contrast, TOAC2 and TOAC13 derivatives presented a single spectral component corresponding to a strong immobilization of the probe. Thus, our findings allowed the description of the peptide topology in the membrane mimetic, where the N-terminal region is in dynamic equilibrium between an ordered, membrane-bound conformation and a disordered, mobile conformation; position 2 is most likely situated in the lipid polar head group region, and residue 13 is fully inserted into the hydrophobic core of the membrane.

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“…As such, many substance classes have been used, including a-and b-amino acids, [32][33][34][35][36][37][38][39] peptoids, [40][41][42] aromatic oligomers, [27,43,44] steroids, [45][46][47] and synthetic polymers. [20,22,23,[48][49][50] Initially, it was believed that the helical rigidity of the AMP backbone ( Figure 1) was a prerequisite for biological activity.…”

Section: Introductionmentioning

confidence: 99%

Synthetic Mimics of Antimicrobial Peptides—A Versatile Ring‐Opening Metathesis Polymerization Based Platform for the Synthesis of Selective Antibacterial and Cell‐Penetrating Polymers

Lienkamp

Tew

2009

Chemistry A European J

147

139

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Natural macromolecules exhibit an extensive arsenal of properties, many of which have proven difficult to recapitulate in simpler synthetic systems. Over the last couple of years, foldamers have emerged as one important step toward increased functionality in synthetic systems. While the great majority of work in this area has focused on folded structures, hence the name, more recent progress has centered on polymers that mimic protein function. These efforts have resulted in the design of relatively simple macromolecules; one example are the synthetic mimics of antimicrobial peptides (SMAMPs) that capture the central physicochemical features of their natural archetypes irrespective of the specific folded form. Here we present our recent efforts to create polymers which display biological activity similar to natural proteins, including antimicrobial and cell-penetrating peptides.

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Combinatorial Synthesis and Directed Evolution Applied to the Production of α-Helix Forming Antimicrobial Peptides Analogues

Cited by 28 publications

References 40 publications

The Designer Antimicrobial Peptide A-hBD-2 Facilitates Skin Wound Healing by Stimulating Keratinocyte Migration and Proliferation

The Designer Antimicrobial Peptide A-hBD-2 Facilitates Skin Wound Healing by Stimulating Keratinocyte Migration and Proliferation

Dynamics and Conformational Studies of TOAC Spin Labeled Analogues of Ctx(Ile21)-Ha Peptide from Hypsiboas albopunctatus

Synthetic Mimics of Antimicrobial Peptides—A Versatile Ring‐Opening Metathesis Polymerization Based Platform for the Synthesis of Selective Antibacterial and Cell‐Penetrating Polymers

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