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Design SMAP29-LysPA26 as a Highly Efficient Artilysin against Pseudomonas aeruginosa with Bactericidal and
Tingting Wang1,2, Yongxiang Zheng1,2, Jiami Dai1,2
1Department of Biopharmaceutics, West China School of Pharmacy, Sichuan University, Chengdu, China.
Abstract:
Antimicrobial resistance (AMR) is a major issue to global health. The multidrug-resistant (MDR) Gram-negative infections, particularly infected by carbapenem-resistant pathogens, urgently need efficient antibiotics and novel therapy. However, the scientific challenges of aiming for innovative approaches against Gram-negative bacteria have hindered the research and development of antibiotic drugs. Phage-derived endolysins are bacteriolytic and specific for a bacterial species or genus, providing a promising antibiotic strategy. However, the outer membrane of Gram-negative bacteria could prevent the peptidoglycan layer from the hydrolysis of endolysins. Antimicrobial peptides usually destabilize the outer membrane and could enhance the antibiotic activity of endolysins. In this study, we designed new artilysins with antimicrobial-peptide SMAP29 fusion at the N-terminal of LysPA26 (named as AL-3AA, AL-9AA, and AL-15AA), and evaluated them. The results showed artilysin AL-3AA to be highly bactericidal; even 0.05 mg/mL AL-3AA could reduce 5.81 log units P. aeruginosa without EDTA in 60 min. It killed P. aeruginosa rapidly and dose-dependently through cell lysis. AL-3AA inhibited P. aeruginosa PAO1 biofilm formation and significantly decreased mature P. aeruginosa biofilms. It also had potential broad-spectrum activity against susceptible Gram-negative bacteria in the hospital, including K. pneumoniae and E. coli. The antibacterial mechanism investigation has provided valuable information about the antibacterial action of AL-3AA, which can lyse and disintegrate the bacterial quickly. These results suggested AL-3AA could be a new and promising antimicrobial agent for the combat of P. aeruginosa. IMPORTANCE Antimicrobial resistance (AMR) is a major issue to global health, particularly the multidrug-resistant (MDR) Gram-negative infections, which pose great challenges. Even new antibiotics research is ongoing, antibiotics used to treat Gram-negative bacteria in the clinical are limited in a small set of molecular scaffolds, and biomolecular categories of antibiotics are urgently needed. In this study, we designed new proteins by combining antimicrobial peptides and endolysins for synergistic bactericidal effects. One of designed proteins, named AL-3AA, showed highly bactericidal, and killed P. aeruginosa rapidly and dose-dependently through cell lysis. It also killed Klebsiella pneumoniae and Escherichia coli, showing potential broad-spectrum activity against susceptible Gram-negative bacteria in the hospital. All results suggest AL-3AA could be a new and promising antimicrobial agent for the combat of P. aeruginosa.
Insights
Antimicrobial resistance (AMR) is a global health threat. Researchers developed artilysin AL-3AA, a novel antimicrobial peptide-endolysin fusion, showing potent bactericidal activity against Pseudomonas aeruginosa and other Gram-negative bacteria.
Area of Science:
- Microbiology
- Biotechnology
- Infectious Diseases
Background:
- Antimicrobial resistance (AMR) poses a significant global health challenge, particularly multidrug-resistant (MDR) Gram-negative infections.
- Developing novel therapies against Gram-negative bacteria is hindered by scientific challenges.
- Phage-derived endolysins offer a promising strategy, but their efficacy is limited by the Gram-negative outer membrane.
Purpose of the Study:
- To design and evaluate novel artilysins, fusion proteins combining antimicrobial peptides and endolysins, for enhanced antibacterial activity.
- To investigate the bactericidal efficacy and mechanism of action of engineered artilysins against Gram-negative pathogens.
- To assess the potential of these novel agents in combating challenging infections like carbapenem-resistant pathogens.
Main Methods:
- Engineered artilysins by fusing the antimicrobial peptide SMAP29 to the N-terminus of LysPA26, creating variants AL-3AA, AL-9AA, and AL-15AA.
- Evaluated the bactericidal activity of artilysins against Pseudomonas aeruginosa, including dose-response and time-kill assays.
- Assessed the effect of artilysin AL-3AA on P. aeruginosa biofilm formation and its activity against other Gram-negative bacteria like Klebsiella pneumoniae and Escherichia coli.
Main Results:
- Artilysin AL-3AA demonstrated potent and rapid bactericidal activity against P. aeruginosa, reducing bacterial load significantly within 60 minutes.
- AL-3AA exhibited dose-dependent killing of P. aeruginosa through cell lysis.
- AL-3AA effectively inhibited P. aeruginosa PAO1 biofilm formation and reduced mature biofilms, also showing activity against K. pneumoniae and E. coli.
Conclusions:
- Artilysin AL-3AA is a highly effective bactericidal agent against P. aeruginosa, offering a promising new therapeutic strategy.
- The engineered artilysin exhibits potential broad-spectrum activity against susceptible Gram-negative bacteria, addressing a critical need in combating AMR.
- The study provides valuable insights into the antibacterial mechanism of AL-3AA, highlighting its potential as a novel antimicrobial agent for clinical applications.
