2021 Fiscal Year Annual Research Report
発電機能を持つ歯周病細菌におけると鉄イオン排出機構に関する研究
| Project/Area Number |
21F21412
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| Research Institution | National Institute for Materials Science |
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Principal Investigator |
岡本 章玄 国立研究開発法人物質・材料研究機構, 国際ナノアーキテクトニクス研究拠点, グループリーダー (70710325)
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| Co-Investigator(Kenkyū-buntansha) |
NARADASU DIVYA 国立研究開発法人物質・材料研究機構, 国際ナノアーキテクトニクス拠点, 外国人特別研究員
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| Project Period (FY) |
2021-11-18 – 2024-03-31
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| Keywords | Extracellula / electron transfer / Membrane vesicles / Redox-repressor rex / Capnocytophaga ochracea / Streptococcus mutans |
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| Outline of Annual Research Achievements |
Bacterial outer membrane vesicles (OMVs) are spherical lipid bilayer nanostructures released by bacteria that facilitate oral biofilm formation via cellular aggregation and intercellular communication. Recent studies have revealed that Capnocytophaga ochracea is one of the dominant members of oral biofilms; however, their potential for OMV production has yet to be investigated. This study demonstrated the biogenesis of OMVs in C. ochracea associated with the concentration of unsaturated fatty acids of phosphatidylinositol (PI) and characterized the size and protein profile of OMVs produced at growth phases. Transmission electron microscopy showed isolated spherical structures from cells stained with heavy metals, indicating the production of OMVs with a size ranging from 25 to 100 nm. Lipidome analysis revealed the presence of phosphatidic acid, phosphatidylethanolamine, phosphatidylcholine, and PI as the main lipids. Some unsaturated fatty acids of PI were present specifically in OMV and little in the outer membrane, suggesting that OMVs are generated from a specific region of the membrane through blebbing rather than a random process such as cell lysis. Furthermore, the lack of similar PI accumulation in the OMV of Porphyromonas gingivalis suggests that C. ochracea has a different biogenesis mechanism. The present study provides a basis for further understanding the roles of C. ochracea OMVs in oral biofilms as well as systemic diseases that C. ochracea involves.
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| Current Status of Research Progress |
Current Status of Research Progress
2: Research has progressed on the whole more than it was originally planned.
Reason
We need to identify the possible electron transfer protein in these bacteria and type of proteins present on the surface of membrane vesicles, which is a challenging part and require interdisciplinary experimental methods and analyses such as transcriptomics, proteomics, gene knock outs and electrochemical characterization.
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| Strategy for Future Research Activity |
I plan to extract the membrane from the pathogens and analyze by mass spectroscopy to identify the proteins, similarly for OMVs.
Isolate the total RNA of pathogens under EET and OCV conditions, to check differentially expressed genes by the transcriptomic analysis.
Biosensor development with a strategy based on substrate specific detection of pathogens.
As new project, in future I test the export of iron either in Fe3+(insoluble) or Fe2+ (soluble) form out of the cell is not well characterized in Porphyromonas gingivalis. Extracellular electron transfer (EET) coupled iron reduction enhanced the biofilm formation and metabolism in a gut pathogen Enterococcus faecalis and in our studies PG has showed the EET capability. Hence, their amino acid based EET can be coupled with the iron reduction implicated in inflammation. Here I propose to research a form of EET coupled iron export induced inflammation in P. gingivalis where, the recycling/export of iron is coupled with the EET to maintain iron homeostasis in the cytoplasm augment its survival causing inflammation on human cells/macrophages by ROS generation. As new approaches for combating bacterial infections and new routes for studying the fundamental mechanisms to be explored which can be applied as critical knowledge for drug discovery for pathogenicity by using electrochemical techniques.
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