2007 Fiscal Year Final Research Report Summary
Elucidation of functional properties of thermostable thermostable cytochromes c and its application to minute molecular design
| Project/Area Number |
17350081
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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 |
Chemistry related to living body
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| Research Institution | University of Tsukuba |
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Principal Investigator |
YAMAMOTO Yasuhiko University of Tsukuba, Graduate School of Pure and Aplied Sciences, Professor (00191453)
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| Co-Investigator(Kenkyū-buntansha) |
NAGATOMO Shigenori University of Tsukuba, Graduate School of Pure and Applied Sciences, Assistant Professor (80373190)
KAWANO Shin University of Tsukuba, Graduate School of Pure and Applied Sciences, Research associate (90431676)
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| Project Period (FY) |
2005 – 2007
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| Keywords | Paramagnetic NMR / Heme comulex / Metalloprotein / Electrochemistry / Coordination bond / Redox potential / Thermostability / Thermodynamics |
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| Research Abstract |
Pseudomonas aeruginosa cytochrome C551 and a series of its mutants exhibiting various thermostabilities have been studied by paramagnetic ^1H NMR and cyclic voltammetry in order to elucidate the molecular mechanisms responsible for control of the redox potentials (E^<o'>) of the proteins. The study revealed that the E^<o'> value of the protein is regulated by two molecular mechanisms operating independently of each other. One is based on the Fe-Met coordination bond strength in the protein, which is determined by the amino acid side-chain packing in the protein, and the other on the pKa value of the heme 17-propionic acid side-chain, which is affected by the electrostatic environment. The former mechanism alters the magnitude of the E^<o'> value throughout the entire pH range and the latter regulates the pK values reflected by the pH profile of the E^<o'> value. These findings provide novel insights into functional regulation of the protein, which could be utilized for tuning the E^<o'> value of the protein by means of protein engineering.
We also succeeded in designing cytochrome c. Thermophile Hydrogenobacter thermophilus cytochrome c552 (HT) is a stable protein with denaturation temperatures ? of 109.8 and 129.7 ℃ for the oxidized and reduced forms, respectively. The removal of a single hydroxyl group from the hydrophobic core of HT, through the replacement of a Tyr by Phe, resulted in further elevation of the Tm value of the oxidized form by 〜6℃, the T_m value of the reduced one remaining essentially unaltered. As a result, the redox potential of the mutant with higher stability in the oxidized form exhibited a negative shift of 〜20 mV relative to that of wild-type HT in an enthalpic manner. These findings indicated that the redox function of a protein can be enthalpically regulated through the stability of the oxidized form by altering the contextual stereochemical packing of hydrophobic residues in the protein interior using protein engineering.
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[Journal Article] Roles of a short connecting disulfide bond in the stability and function of psychrophilic Sbewanella violacea cytochrome c52007
Author(s)
K. Ogawa, T. Sonoyama, T. Takeda, S. Ichiki, S. Nakamura, Y. Kobayashi, S. Uchiyama, K. Nakasone, S. J. Takayama, H. Mita, Y. Yamamoto, and Y. Sambongi
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Journal Title
Extremophiles 11
Pages: 797-807
Description
「研究成果報告書概要(欧文)」より
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[Presentation] Characterization of electron transfer from cytochrome cto blue copper proteins2009
Author(s)
S. J. Takayama, H. Tai, S. Mikami, K. Irie, M. Kage, S. Kawano, S, Nagatomo, T. Kawahara, N. Funasaki, A. Takabe, S. Hirota, and Y. Yamamoto
Organizer
Global COE in Chemistry, Nagoya Special Symposium
Place of Presentation
Nagoya Univ
Year and Date
2009-03-21
Description
「研究成果報告書概要(欧文)」より
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[Presentation] Electron transfer from cytochrome c to blue copper Proteins2008
Author(s)
K. Irie, S. J. Takayama, H. Tai, S. Mikami, M. Kage, S. Kawano, S, Nagatomo, T. Kawahara, N. Funasaki, A. Takabe, S. Hirota, and Y. Yamamoto
Organizer
First International Symposium on Interdisciplinary Materials Science
Place of Presentation
Tsukuba City
Year and Date
2008-03-13
Description
「研究成果報告書概要(欧文)」より
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[Presentation] Reorganization energy of electron transfer from cytochrome c to blue copper proteins2007
Author(s)
S. J. Takayama, H. Tai, S. Mikami, K. Irie, M. Kage, S. Kawano, T. Kawahara, N. Funasaki, A. Takabe, S. Hirota, and Y. Yamamoto
Organizer
13th International Conference on Biological Inorganic Chemistry
Place of Presentation
Vienna, Austria
Year and Date
2007-07-18
Description
「研究成果報告書概要(欧文)」より
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[Presentation] High pressure NMR studies on internal dynamics of redox active protein cytochrome c2006
Author(s)
S. J. Takayama, Y. Takahashi, S. Mikami, H. Tai, S. Kawano, H. Mita, Y. Yamamoto, Y. Sambongi, R. Kitahara, S. Yokoyama, and K. Akasaka
Organizer
55th Annual Meeting of Polymer Society of Japan
Place of Presentation
Toyama Univ
Year and Date
2006-09-21
Description
「研究成果報告書概要(欧文)」より
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[Presentation] Characterization of internal dynamics of thermophile Hydrogenobacter thermophilus cytochrome c Using High Pressure NMR2006
Author(s)
S. J. Takayama, Y. Takahashi, S. Mikami, S. Nagatomo, H. Mita, Y. Yamamoto, Y. Sambongi, R. Kitahara, S. Yokoyama, and K. Akasaka
Organizer
XXIIND International Conference on Magnetic Resonance in Biological Systems
Place of Presentation
Gottingen, German
Year and Date
2006-08-22
Description
「研究成果報告書概要(欧文)」より
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[Presentation] High Pressure NMR Study of Thermophile Hydrogenobacter thermophilus Cytochrome c5522006
Author(s)
S. J. Takayama, Y. Takahashi, H. Tai, S. Mikami, H. Sasaki, S. Hirota, H. Mita, Y. Yamamoto, Y. Sambongi, H. Hemmi, R. Kitahara, S. Yokoyama, and K. Akasaka
Organizer
47th Experimental Nuclear Magnetic Resonance Conference
Place of Presentation
Pacific Grove, California, USA
Year and Date
2006-04-24
Description
「研究成果報告書概要(欧文)」より
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