Semi-Biosynthetic Production of Surface-Binding Adhesive Antimicrobial Peptides Using Intein-Mediated Protein Ligation

Hwang, Young Eun; Im, Sang Hee; Cho, Ju Hyun; Lee, Wonsik; Cho, Byung-Kwan; Sung, Bong Hyun; Kim, Sun Chang

doi:10.3390/ijms232315202

Cited by 3 publications

(1 citation statement)

References 48 publications

(51 reference statements)

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“…These peptides have a wide range of activities against Gram-negative and -positive bacteria, fungi, parasites, and viruses. AMP production can be performed by chemical synthesis ,− or using recombinant expression systems. − AMP synthesis also aims to achieve greater selectivity and a decrease in hemolytic activity or cytotoxicity for the host cells.…”

Section: Introductionmentioning

confidence: 99%

Anti-Staphy Peptides Rationally Designed from Cry10Aa Bacterial Protein

Rios,

Maximiano,

Fernandes

et al. 2024

ACS Omega

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Bacterial infections pose a significant threat to human health, constituting a major challenge for healthcare systems. Antibiotic resistance is particularly concerning in the context of treating staphylococcal infections. In addressing this challenge, antimicrobial peptides (AMPs), characterized by their hydrophobic and cationic properties, unique mechanism of action, and remarkable bactericidal and immunomodulatory capabilities, emerge as promising alternatives to conventional antibiotics for tackling bacterial multidrug resistance. This study focuses on the Cry10Aa protein as a template for generating AMPs due to its membrane-penetrating ability. Leveraging the Joker algorithm, six peptide variants were derived from α-helix 3 of Cry10Aa, known for its interaction with lipid bilayers. In vitro, antimicrobial assays determined the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) required for inhibiting the growth of Staphylococcus aureus, Escherichia coli, Acinetobacter baummanii, Enterobacter cloacae, Enterococcus facallis, Klebsiella pneumonia, and Pseudomonas aeruginosa. Time-kill kinetics were performed using the parental peptide AMPCry10Aa, as well as AMPCry10Aa_1 and AMPCry10Aa_5, against E. coli ATCC, S. aureus 111 and S. aureus ATCC strains showing that AMPCry10Aa_1 and AMPCry10Aa_5 peptides can completely reduce the initial bacterial load with less than 2 h of incubation. AMPCry10Aa_1 and AMPCry 10Aa_5 present stability in human serum and activity maintenance up to 37 °C. Cytotoxicity assays, conducted using the MTT method, revealed that all of the tested peptides exhibited cell viability >50% (IC50). The study also encompassed evaluations of the structure and physical-chemical properties. The three-dimensional structures of AMPCry10Aa and AMPCry10Aa_5 were determined through nuclear magnetic resonance (NMR) spectroscopy, indicating the adoption of α-helical segments. Electron paramagnetic resonance (EPR) spectroscopy elucidated the mechanism of action, demonstrating that AMPCry10Aa_5 enters the outer membranes of E. coli and S. aureus, causing substantial increases in lipid fluidity, while AMPCry10Aa slightly increases lipid fluidity in E. coli. In conclusion, the results obtained underscore the potential of Cry10Aa as a source for developing antimicrobial peptides as alternatives to conventional antibiotics, offering a promising avenue in the battle against antibiotic resistance.

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Section: Introductionmentioning

confidence: 99%

Anti-Staphy Peptides Rationally Designed from Cry10Aa Bacterial Protein

Rios,

Maximiano,

Fernandes

et al. 2024

ACS Omega

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A systematical review on antimicrobial peptides and their food applications

Xu,

Zhao,

Tan

et al. 2023

Biomaterials Advances

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Design of a recombinant asparaginyl ligase for site-specific modification using efficient recognition and nucleophile motifs

Tang,

Hao,

Liu

et al. 2024

Commun Chem

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Asparaginyl ligases have been extensively utilized as valuable tools for site-specific bioconjugation or surface-modification. However, the application is hindered by the laborious and poorly reproducible preparation processes, unstable activity and ambiguous substrate requirements. To address these limitations, this study employed a structure-based rational approach to obtain a high-yield and high-activity protein ligase called OaAEP1-C247A-aa55-351. It was observed that OaAEP1-C247A-aa55-351 exhibits appreciable catalytic activities across a wide pH range, and the addition of the Fe3+ metal ion effectively enhances the catalytic power. Importantly, this study provides insight into the recognition and nucleophile peptide profiles of OaAEP1-C247A-aa55-351. The ligase demonstrates a higher recognition ability for the “Asn-Ala-Leu” motif and an N-terminus “Arg-Leu” as nucleophiles, which significantly increases the reaction yield. Consequently, the catalytic activity of OaAEP1-C247A-aa55-351 with highly efficient recognition and nucleophile motif, “Asn-Ala-Leu” and “Arg-Leu” under the buffer containing Fe3+ is 70-fold and 2-fold higher than previously reported OaAEP1-C247A and the most efficient butelase-1, respectively. Thus, the designed OaAEP1-C247A-aa55-351, with its highly efficient recognition and alternative nucleophile options, holds promising potential for applications in protein engineering, chemo-enzymatic modification, and the development of drugs.

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Semi-Biosynthetic Production of Surface-Binding Adhesive Antimicrobial Peptides Using Intein-Mediated Protein Ligation

Cited by 3 publications

References 48 publications

Anti-Staphy Peptides Rationally Designed from Cry10Aa Bacterial Protein

Anti-Staphy Peptides Rationally Designed from Cry10Aa Bacterial Protein

A systematical review on antimicrobial peptides and their food applications

Design of a recombinant asparaginyl ligase for site-specific modification using efficient recognition and nucleophile motifs

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