2015 Fiscal Year Annual Research Report
過飽和の解消による蛋白質の異常凝集体の形成に関する研究
| Project/Area Number |
14J04433
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| Research Institution | Osaka University |
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Principal Investigator |
林 雨曦 大阪大学, 理学研究科, 特別研究員(DC1)
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| Project Period (FY) |
2014-04-25 – 2017-03-31
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| Keywords | cytochrome c / protein aggregation / solubility / supersaturation / phase diagram |
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| Outline of Annual Research Achievements |
This year I successfully finished the research project of cytochrome c (cyt c) aggregation (Lin et al. (2016) Langmuir 32, 2010-2022). I examined the aggregation behavior of the physiologically relevant two types of cyt c, metal-bound cyt c, and its fragment with high amyloidogenicity in alcohol/water mixtures. The aggregation propensity of holo cyt c was low due to high solubility, and markedly unfolded apo cyt c, lacking the heme prosthetic group, strongly promoted the propensity for amorphous aggregation with increases in hydrophobicity. Silver-bound apo cyt c increased the capacity of fibrillar aggregation due to subtle structural changes of apo cyt c by strong binding of silver. However, mature amyloid fibrils were not detected for any of the cyt c variants or its fragment, even with extensive ultrasonication. These results revealed the intrinsically low amyloidogenicity of cyt c, which is beneficial for its homeostasis and function by facilitating the folding and minimizing irreversible amyloid formation. I proposed that intrinsically low amyloidogenicity of cyt c is attributed to the low metastability of supersaturation. I also demonstrated that the phase diagram constructed based on solubility and aggregate type is useful for a comprehensive understanding of protein aggregation. Furthermore, amorphous aggregation, which is also viewed as a generic property of proteins, and amyloid fibrillation can be distinguished from each other by the metastability of supersaturation.
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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
The phase diagram of cytochrome c has been constructed as the research plan. This phase diagram revealed the intrinsically low amyloidogenicity of cytochrome c and that the balance between amorphous and fibrillar aggregates depends on stability of supersaturated solutions of this protein (Langmuir 2016).
I have also extended my study on protein aggregation to a more biologically relevant system by means of vesicles mimicking biological membranes. The effect of vesicles on alpha-synuclein fibrillation has been examined.
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| Strategy for Future Research Activity |
The effects of small molecules on vesicle-dependent fibrillation of alpha-synuclein fibrillation will be examined using various biophysical techniques. Based on the experimental data, the phase diagram of alpha-synuclein will be constructed and the effect of small molecules on alpha-synuclein fibrillation will be evaluated from the point of view of solubility and supersaturation.
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