2011 Fiscal Year Final Research Report
Survival strategy of intestinal lactic acid bacteria in the gut : functional analysis of cell surface structure involved in bile acid adaptation
| Project/Area Number |
21380053
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Applied microbiology
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| Research Institution | Hokkaido University |
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Principal Investigator |
YOKOTA Atsushi 北海道大学, 大学院・農学研究院, 教授 (50220554)
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| Co-Investigator(Kenkyū-buntansha) |
FUKIYA Satoru 北海道大学, 大学院・農学研究院, 助教 (10370157)
MORITA Naoki 独立行政法人産業技術総合研究所, 生物プロセス研究部門, 研究グループ長 (60371085)
YOKOTA Shin-ichishi 札幌医科大学, 医学部, 准教授 (10325863)
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| Project Period (FY) |
2009 – 2011
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| Keywords | 微生物機能 / 乳酸菌 |
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| Research Abstract |
Free bile acids (FBAs) are considered the most deleterious stress compounds that inhibit the growth of intestinal bacteria by their membrane-damaging effect. Previously, we found adaptation in Lactobacillus gasseri JCM1131T to cholic acid (CA), one of the major FBAs in human intestine. In this adaptation, exponentially growing cells were first exposed to 4 mM CA, a sub-lethal concentration, for 30 min and then to lethal concentration at 15 mM. The adapted cells did not show any appreciable decrease in viability in the presence of 15 mM CA even after 7 h, while non-adapted cells rapidly lost viability by the factors of 10-3~-4. In the adapted cells a significant relief of membrane damage was observed in the presence of CA, suggesting alterations in lipid composition in the cell envelope.
Total lipids extracted from adapted and non-adapted cells were separated into three fractions, neutral lipids, glycolipids and phospholipids. Determination of amount of each fraction revealed no signific
… More
ant changes between adapted and non-adapted cells. Likewise, significant differences were neither found in fatty acids compositions of each fraction. However, significant alterations were detected in glycolipids and phospholipids compositions. In glycolipids, increases in the length of sugar chain were detected during the adaptation. Furthermore, a significant increase in relative amount of cardiolipin (CL) was observed in the phospholipid fraction.
To evaluate physiological importance of CL in CA adaptation, vesicle experiment was conducted. It was found that CL confers resistance to the liposome against CA attack, suggesting a pivotal role of CL in adaptation mechanism.
Mutants having deletion in two putative cardiolipin synthase genes (cls) singly or in combination were then derived, and their abilities in CA adaptation were investigated. Although CL levels of the both single-knockout mutants were lower than that of the wild type JCM1131T, the levels were found to increase after exposure to 4 mM CA, and a comparable CA adaptation to the wild type strain was observed in both the mutants. The double-knockout mutant possessed negligible amount of CL but still showed CA adaptation, suggesting a dispensable role of CL in CA adaptation. However, in this mutant, significant decrease in glycolipid fraction and significant increase in phospholipid fraction were observed as compared to wild-type strain and these shifts were more prominent after CA adaptation. Therefore, CL was found to work as a key determinant for the lipid composition of the bacterial cell membrane. Less
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