Addition of L-cysteine to the N- or C-terminus of the all-D-enantiomer [D(KLAKLAK)2] increases antimicrobial activities against multidrug-resistant Pseudomonas aeruginosa, Acinetobacter baumannii and Escherichia coli

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Addition of L-cysteine to the N- or C-terminus of the all-D-enantiomer [D(KLAKLAK)2] increases antimicrobial activities against multidrug-resistant Pseudomonas aeruginosa, Acinetobacter baumannii and Escherichia coli https://t.co/zdXvQD54g1 @thePeerJ https://t.co/Mw4zYesx21
Addition of L-cysteine to the N- or C-terminus of the all-D-enantiomer increases antimicrobial activities against multidrug-resistant Pseudomonas aeruginosa, Acinetobacter baumannii and Escherichia coli @thePeerJ article https://t.co/sMrUmGsS0E #Biochemistry #Microbiology https://t.co/xcwvOujKZY
Microbiology

Main article text

 

Introduction

Materials & Methods

Peptide design and synthesis

Bacterial strains

Drug susceptibility testing

Cytotoxicity tests

Statistical analysis

Results

Addition of L-cysteine to the N- or C- terminus enhanced the antimicrobial activity of the original peptide

Synergistic effects of peptides and antibiotics

Cytotoxicity of antimicrobial peptides to HepG2 cells

Discussion

Conclusions

Supplemental Information

HPLC chromatogram and MALDI-TOF mass spectrum of DP

(A) Chromatogram of DP using reversed phase HPLC. DP was loaded onto a SunFire C18 column and eluted with a linear acetonitrile gradient (10–60%) in 20 min. (B) Mass spectra of DP.

DOI: 10.7717/peerj.10176/supp-1

HPLC chromatogram and MALDI-TOF mass spectrum of C-DP

(A) Chromatogram of C-DP using reversed phase HPLC. C-DP was loaded onto a SunFire C18 column and eluted with a linear acetonitrile gradient (10–60%) in 20 min. (B) Mass spectra of C-DP.

DOI: 10.7717/peerj.10176/supp-2

HPLC chromatogram and MALDI-TOF mass spectrum of DP-C

(A) Chromatogram of DP-C using reversed phase HPLC. DP-C was loaded onto a SunFire C18 column and eluted with a linear acetonitrile gradient (0–100%) in 20 min. (B) Mass spectra of DP-C.

DOI: 10.7717/peerj.10176/supp-3

HPLC chromatogram and MALDI-TOF mass spectrum of DP-C dimer

(A) Chromatogram of DP-C dimer using reversed phase HPLC. DP-C dimer was loaded onto a COSMOSIL 5C18-AR-300 column and eluted with a linear acetonitrile gradient (20–50%) in 20 min. (B) Mass spectra of DP-C dimer.

DOI: 10.7717/peerj.10176/supp-4

Cytotoxicity of DP, C-DP or DP-C on HepG2 cell lines

HepG2 cells were seeded at 3,000 cells/well in 96-well microtiter plates. After incubation for 48 h, DP (0–256 µg/ml), C-DP (0–256 µg/ml) or DP-C (0–256 µg/ml) were added, then the plates incubated for an additional 48 h. Cell viability was determined using a Cell Counting Kit-8, with colorimetric changes assessed at OD450 nm with a microplate reader.

DOI: 10.7717/peerj.10176/supp-5

Cytotoxicity of high-dose colistin on HepG2 cell lines

HepG2 cells were seeded at 3,000 cells/well in 96-well microtiter plates. After incubation for 48 h, colistin (0–3,000 µg/ml) were added, then the plates incubated for an additional 48 h. Cell viability was determined using a Cell Counting Kit-8, with colorimetric changes assessed at OD450 nm with a microplate reader.

DOI: 10.7717/peerj.10176/supp-6

Figure 1 raw data: Cuytotoxicity assay of DP-C and colistin

DOI: 10.7717/peerj.10176/supp-7

Figure S1 raw data: Cytotoxicity assay of peptides

DOI: 10.7717/peerj.10176/supp-8

Figure S2 raw data: Cytotoxicity assay of colistin

DOI: 10.7717/peerj.10176/supp-9

Additional Information and Declarations

Competing Interests

The authors declare there are no competing interests.

Author Contributions

Maki K. Ohno and Isao Ishida conceived and designed the experiments, performed the experiments, analyzed the data, prepared figures and/or tables, authored or reviewed drafts of the paper, and approved the final draft.

Teruo Kirikae analyzed the data, prepared figures and/or tables, authored or reviewed drafts of the paper, and approved the final draft.

Eisaku Yoshihara analyzed the data, authored or reviewed drafts of the paper, and approved the final draft.

Fumiko Kirikae performed the experiments, prepared figures and/or tables, and approved the final draft.

Data Availability

The following information was supplied regarding data availability:

The raw measurements are available in Supplemental Files.

Funding

This investigation was supported by Grant-in-Aid for Scientific Research (C26460173), MEXT (Ministry of Education, Culture, Sports, Science and Technology)-Supported Program for the Strategic Research Foundation at Private Universities, 2014-2018 and a grant from the Naito Foundation. There was no additional external funding received for this study. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

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