Rational design of crystal contact-free space in protein crystals for analyzing spatial distribution of motions within protein molecules

2018 Fiscal Year Final Research Report

• Project Area: Novel measurement techniques for visualizing 'live' protein molecules at work
• Project/Area Number: 26119002
• Research Category: Grant-in-Aid for Scientific Research on Innovative Areas (Research in a proposed research area)
• Allocation Type: Single-year Grants
• Review Section: Complex systems
• Research Institution: Kyushu University
• Principal Investigator: Kohda Daisuke 九州大学, 生体防御医学研究所, 教授 (80186618)
• Co-Investigator(Kenkyū-buntansha): 稲垣 冬彦 公益財団法人微生物化学研究会, 微生物化学研究所, 客員研究員 (70011757)
• Project Period (FY): 2014-07-10 – 2019-03-31
• Keywords: タンパク質結晶 / 動的構造 / 結晶コンタクト / 融合タンパク質 / MBP / Tom20 / Tim21

Outline of Final Research Achievements

We developed a fusion protein method to create crystal contact-free space (CCFS) in protein crystals and to place the mobile parts or ligands in the CCFS. The mobile parts/ligands appear as smeared electron densities in the CCFS in the difference electron density map. We applied the CCFS method to visualize the movement of a highly mobile presequence peptide as bound to the mitochondrial import receptor, Tom20, and to estimate the solution conformation of a flexible loop segment in another mitochondrial import protein, Tim21, which was distorted in the conventional crystals by the crystal contacts.

Free Research Field

構造生物学

Academic Significance and Societal Importance of the Research Achievements

蛋白質の結晶中では分子同士が接触して３次元の結晶格子を作っている．蛋白質の中には一部の構造が柔らかく創られているために，結晶中では結晶コンタクトにより容易に変形し，正しい形（溶液中の平衡状態）をとっていない可能性がある．柔らかく動的な部分の構造変化は蛋白質の機能と密接に関連していることが少なくないことを考えると，「柔動構造の変形・固定問題」を真剣にとらえ，実験的に解決する手段を考案することは重要である．