Single molecule resolution of the antimicrobial action of quantum dot-labeled sushi peptide on live bacteria
Background: Antimicrobial peptides are found in all kingdoms of life. During the evolution of multicellular organisms, antimicrobial peptides were established as key elements of innate immunity. Most antimicrobial peptides are thought to work by disrupting the integrity of cell membranes, causing pa...
| Main Authors: | , , , , , |
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| Other Authors: | |
| Format: | Article |
| Language: | en_US |
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BioMed Central
2010
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| Online Access: | http://hdl.handle.net/1721.1/52522 https://orcid.org/0000-0002-5687-6154 |
| _version_ | 1811085242635649024 |
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| author | Ding, Jeak Ling Wohland, Thorsten Ho, Bow Har, Jia Yi Leptihn, Sebastian Chen, Jianzhu |
| author2 | Massachusetts Institute of Technology. Department of Biology |
| author_facet | Massachusetts Institute of Technology. Department of Biology Ding, Jeak Ling Wohland, Thorsten Ho, Bow Har, Jia Yi Leptihn, Sebastian Chen, Jianzhu |
| author_sort | Ding, Jeak Ling |
| collection | MIT |
| description | Background: Antimicrobial peptides are found in all kingdoms of life. During the evolution of multicellular organisms, antimicrobial peptides were established as key elements of innate immunity. Most antimicrobial peptides are thought to work by disrupting the integrity of cell membranes, causing pathogen death. As antimicrobial peptides target the membrane structure, pathogens can only acquire resistance by a fundamental change in membrane composition. Hence, the evolution of pathogen resistance has been a slow process. Therefore antimicrobial peptides are valuable alternatives to classical antibiotics against which multiple drug-resistant bacteria have emerged. For potential therapeutic applications as antibiotics a thorough knowledge of their mechanism of action is essential. Despite the increasingly comprehensive understanding of the biochemical properties of these peptides, the actual mechanism by which antimicrobial peptides lyse microbes is controversial.
Results: Here we investigate how Sushi 1, an antimicrobial peptide derived from the horseshoe crab (Carcinoscorpius rotundicauda), induces lysis of Gram-negative bacteria. To follow the entire process of antimicrobial action, we performed a variety of experiments including transmission electron microscopy and fluorescence correlation spectroscopy as well as single molecule tracking of quantum dot-labeled antimicrobial peptides on live bacteria. Since in vitro measurements do not necessarily correlate with the in vivo action of a peptide we developed a novel fluorescent live bacteria lysis assay. Using fully functional nanoparticle-labeled Sushi 1, we observed the process of antimicrobial action at the single-molecule level.
Conclusion: Recently the hypothesis that many antimicrobial peptides act on internal targets to kill the bacterium has been discussed. Here, we demonstrate that the target sites of Sushi 1 are outer and inner membranes and are not cytosolic. Further, our findings suggest four successive steps of the bactericidal process: 1) Binding, mediated mainly by charged residues in the peptide; 2) Peptide association, as peptide concentration increases evidenced by a change in diffusive behavior; 3) Membrane disruption, during which lipopolysaccharide is not released; and 4) Lysis, by leakage of cytosolic content through large membrane defects. |
| first_indexed | 2024-09-23T13:05:51Z |
| format | Article |
| id | mit-1721.1/52522 |
| institution | Massachusetts Institute of Technology |
| language | en_US |
| last_indexed | 2024-09-23T13:05:51Z |
| publishDate | 2010 |
| publisher | BioMed Central |
| record_format | dspace |
| spelling | mit-1721.1/525222022-10-01T13:00:38Z Single molecule resolution of the antimicrobial action of quantum dot-labeled sushi peptide on live bacteria Ding, Jeak Ling Wohland, Thorsten Ho, Bow Har, Jia Yi Leptihn, Sebastian Chen, Jianzhu Massachusetts Institute of Technology. Department of Biology Koch Institute for Integrative Cancer Research at MIT Chen, Jianzhu Chen, Jianzhu Background: Antimicrobial peptides are found in all kingdoms of life. During the evolution of multicellular organisms, antimicrobial peptides were established as key elements of innate immunity. Most antimicrobial peptides are thought to work by disrupting the integrity of cell membranes, causing pathogen death. As antimicrobial peptides target the membrane structure, pathogens can only acquire resistance by a fundamental change in membrane composition. Hence, the evolution of pathogen resistance has been a slow process. Therefore antimicrobial peptides are valuable alternatives to classical antibiotics against which multiple drug-resistant bacteria have emerged. For potential therapeutic applications as antibiotics a thorough knowledge of their mechanism of action is essential. Despite the increasingly comprehensive understanding of the biochemical properties of these peptides, the actual mechanism by which antimicrobial peptides lyse microbes is controversial. Results: Here we investigate how Sushi 1, an antimicrobial peptide derived from the horseshoe crab (Carcinoscorpius rotundicauda), induces lysis of Gram-negative bacteria. To follow the entire process of antimicrobial action, we performed a variety of experiments including transmission electron microscopy and fluorescence correlation spectroscopy as well as single molecule tracking of quantum dot-labeled antimicrobial peptides on live bacteria. Since in vitro measurements do not necessarily correlate with the in vivo action of a peptide we developed a novel fluorescent live bacteria lysis assay. Using fully functional nanoparticle-labeled Sushi 1, we observed the process of antimicrobial action at the single-molecule level. Conclusion: Recently the hypothesis that many antimicrobial peptides act on internal targets to kill the bacterium has been discussed. Here, we demonstrate that the target sites of Sushi 1 are outer and inner membranes and are not cytosolic. Further, our findings suggest four successive steps of the bactericidal process: 1) Binding, mediated mainly by charged residues in the peptide; 2) Peptide association, as peptide concentration increases evidenced by a change in diffusive behavior; 3) Membrane disruption, during which lipopolysaccharide is not released; and 4) Lysis, by leakage of cytosolic content through large membrane defects. Singapore-MIT Alliance (SMA II, Computation and Systems Biology, FRP: Immunology & Tissue Defense) Singapore Bioimaging Consortium (SBIC 003/2005: R-143-000-284-305) 2010-03-11T19:57:52Z 2010-03-11T19:57:52Z 2009-05 2008-11 Article http://purl.org/eprint/type/JournalArticle 1741-7007 http://hdl.handle.net/1721.1/52522 Leptihn, Sebastian et al. “Single molecule resolution of the antimicrobial action of quantum dot-labeled sushi peptide on live bacteria.” BMC Biology 7.1 (2009): 22. https://orcid.org/0000-0002-5687-6154 en_US http://dx.doi.org/10.1186/1741-7007-7-22 BMC Biology Creative Commons Attribution http://creativecommons.org/licenses/by/2.0/ application/pdf BioMed Central BioMed Central |
| spellingShingle | Ding, Jeak Ling Wohland, Thorsten Ho, Bow Har, Jia Yi Leptihn, Sebastian Chen, Jianzhu Single molecule resolution of the antimicrobial action of quantum dot-labeled sushi peptide on live bacteria |
| title | Single molecule resolution of the antimicrobial action of quantum dot-labeled sushi peptide on live bacteria |
| title_full | Single molecule resolution of the antimicrobial action of quantum dot-labeled sushi peptide on live bacteria |
| title_fullStr | Single molecule resolution of the antimicrobial action of quantum dot-labeled sushi peptide on live bacteria |
| title_full_unstemmed | Single molecule resolution of the antimicrobial action of quantum dot-labeled sushi peptide on live bacteria |
| title_short | Single molecule resolution of the antimicrobial action of quantum dot-labeled sushi peptide on live bacteria |
| title_sort | single molecule resolution of the antimicrobial action of quantum dot labeled sushi peptide on live bacteria |
| url | http://hdl.handle.net/1721.1/52522 https://orcid.org/0000-0002-5687-6154 |
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