Targeting of the Essential acpP, ftsZ, and rne Genes in Carbapenem-Resistant Acinetobacter baumannii by Antisense PNA Precision Antibacterials
Abstract
:1. Introduction
2. Materials and Methods
2.1. Bacterial Strains
2.2. Antibiotic MIC
2.3. Computational Screening of Sequence Regions in the mRNA of Target Genes
2.4. BPP-PNA Synthesis
2.5. BPP-PNA MIC
2.6. Time-Kill Assay
2.7. HepG2 Cell Toxicity
2.8. Envelope Disruption
2.9. Real-Time RT-PCR
2.10. Statistics
3. Results
3.1. PNA Design for the mRNA of Target Genes
3.1.1. AcpP
3.1.2. FtsZ
3.1.3. RNase E
3.2. MIC and Time Kill Kinetics
3.3. Species-Selective Activity of BPP-PNAs
3.4. Cytotoxicity
3.5. Envelope Disruption
3.6. Effect on Target mRNA
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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PNA No. | Sequence N→C | Target Position | Peptide | MIC (µM) | ||||
---|---|---|---|---|---|---|---|---|
ATCC 19606 | AC44 | AC46 | SSI 1104 | AC2 | ||||
5652 (m) * | ATATCGCTCAC | +40 to +50 b | H-(KFF)3K-eg1 | 4 | 4 | 4 | 4 | 4 |
5653 (mm) * | ATACCGCTTAC | H-(KFF)3K-eg1 | 4 | 4 | 4 | 4 | 4 | |
5639 (m) * | TGATTTGCCAC | +1 to +11 a | H-(KFF)3K-eg1 | 2 | 1 | 1 | 2 | 1–2 |
5640 (mm) * | TGACTTGTCAC | H-(KFF)3K-eg1 | 4 | 4 | 4 | 4 | 4 | |
5639 (m) ** | TGATTTGCCAC | +1 to +11 a | H-(KFF)3K-eg1 | 4 | 4 | 4 | 4 | 4 |
5640 (mm) ** | TGACTTGTCAC | H-(KFF)3K-eg1 | 4 | 4 | 4 | 4 | 4 | |
5823 (m) * | TGATTTGCCAC | +1 to +11 a | H-(kff)3k-eg1 c | 4 | 4 | 4 | 4 | 4 |
5824 (mm) * | TGACTTGTCAC | H-(kff)3k-eg1 c | 8 | 4 | 4 | 8 | 8 |
PNA No | Sequence N→C | Target Position | Peptide | MIC (µM) | ||
---|---|---|---|---|---|---|
ATCC 19606 | AC44 | SSI1104 | ||||
5612 (m) | GAGGCCATGAC | −3 to +8 | H-(KFF)3K-eg1 | 4 | 8 | 4 |
5615 (mm) | GAGTCCAGGAC | H-(KFF)3K-eg1 | 8 | 16 | 16 | |
5613 (m) | GAGGCCATGAC | −3 to +8 | H-(R-Ahx-R)4-Ahx | >16 | >16 | nd |
5616 (mm) | GAGTCCAGGAC | H-(R-Ahx-R)4-Ahx | >16 | >16 | nd | |
5657 (m) | ATGAGGCCATG | −1 to +10 | H-(KFF)3K-eg1 | >16 | 16 | nd |
5658 (m) | GGCCATGACCT | −5 to +6 | H-(KFF)3K-eg1 | 4 | 8 | 8 |
Strains | H-(KFF)3K-eg1-ftsZ PNA(5612) | H-(KFF)3K-eg1-ftsZ PNA(5615) |
---|---|---|
ATCC19606 | 4 µM | 8 µM |
AC1 | 4 µM | 16 µM |
AC2 | 2 µM | 16 µM |
AC3 | 4 µM | 8 µM |
AC4 | 8 µM | 16 µM |
AC5 | 4 µM | 16 µM |
AC6 | 4 µM | 8 µM |
AC7 | 4 µM | 16 µM |
AC8 | 8 µM | 16 µM |
AC9 | 2 µM | 16 µM |
AC44 | 8 µM | 16 µM |
AC46 | 8 µM | 16 µM |
SSI1104 | 4 µM | 16 µM |
MIC50 | 4 µM | 16 µM |
PNA No | Sequence N→C | Target Position | Peptide | MIC (µM) | ||||
---|---|---|---|---|---|---|---|---|
ATCC 19606 | AC44 | AC46 | SSI 1104 | AC2 | ||||
5637 (m) | ACGTTTCATGG | −2 to +9 | H-(KFF)3K-eg1 | 1 | 1 | 1 | nd | nd |
5638 (mm) | ACGATTCTTGG | H-(KFF)3K-eg1 | 2 | 2 | 1 | nd | nd | |
5654 (m) | ATACGTTTCAT | +1 to +11 | H-(KFF)3K-eg1 | 2 | 1 | 2 | 2 | 2 |
5821 (mm) | ATCCGTTTAAT | H-(KFF)3K-eg1 | 4 | 1 | 2 | 2 | 2 | |
5656 (m) | TTTCATGGGTG | −5 to +6 | H-(KFF)3K-eg1 | 2 | 2 | 2 | 2 | 2 |
5822 (mm) | TTTGATGTGTG | H-(KFF)3K-eg1 | 8 | 8 | 8 | 8 | 8 | |
5655 (m) | GTTTCATGGGT | −4 to +7 | H-(KFF)3K-eg1 | 4 | 8 | nd | nd | 8 |
Target | PNA | Sequence * | MIC (µM) | MIC (µM) | MIC (µM) | MIC (µM) |
---|---|---|---|---|---|---|
P. aeruginosa (PAO1) | E. coli K-12 MG1655 | A. baumannii ATCC 19606 | A. baumannii SSI1104 | |||
ftsZ (m) | 5612 | GAGGCCATGAC | >16 µM | >16 µM | 4 µM | 4 µM |
acpP (m) | 5639 | TGATTTGCCAC | >16 µM | >16 µM | 2 µM | 2 µM |
acpP (m) | 5652 | ATATCGCTCAC | 16 µM | 16 µM | 4 µM | 4 µM |
ftsZ (mm) # | 3965 | TTCTAACAAAGT | nd | 16 µM | 16 µM | ≥16 µM |
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Nejad, A.J.; Shahrokhi, N.; Nielsen, P.E. Targeting of the Essential acpP, ftsZ, and rne Genes in Carbapenem-Resistant Acinetobacter baumannii by Antisense PNA Precision Antibacterials. Biomedicines 2021, 9, 429. https://0-doi-org.brum.beds.ac.uk/10.3390/biomedicines9040429
Nejad AJ, Shahrokhi N, Nielsen PE. Targeting of the Essential acpP, ftsZ, and rne Genes in Carbapenem-Resistant Acinetobacter baumannii by Antisense PNA Precision Antibacterials. Biomedicines. 2021; 9(4):429. https://0-doi-org.brum.beds.ac.uk/10.3390/biomedicines9040429
Chicago/Turabian StyleNejad, Alireza Japoni, Nader Shahrokhi, and Peter E. Nielsen. 2021. "Targeting of the Essential acpP, ftsZ, and rne Genes in Carbapenem-Resistant Acinetobacter baumannii by Antisense PNA Precision Antibacterials" Biomedicines 9, no. 4: 429. https://0-doi-org.brum.beds.ac.uk/10.3390/biomedicines9040429