Development of new NMR method for studying structural dynamics of large molecular proteins

2017 Fiscal Year Final Research Report

• Project/Area Number: 15K06966
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Structural biochemistry
• Research Institution: Osaka University (2017); Nagoya University (2015-2016)
• Principal Investigator: Miyanoiri Yohei 大阪大学, たんぱく質研究所, 准教授 (80547521)
• Project Period (FY): 2015-04-01 – 2018-03-31
• Keywords: NMR / 蛋白質動態 / 安定同位体標識 / 高分子量蛋白質 / 圧力可変実験

Outline of Final Research Achievements

We developed new relaxation optimized SAIL amino acids for studying structural dynamics of large molecular proteins. Moreover, we also developed cost effective E. coli cellular expression systems, using auxotrophic mutants lacking certain enzymes in the biosynthetic pathways of various amino acids. From these results, we could successfully apply them to the 82 kDa MSG and about 1 MDa GroEL-GroES to observe the 13CH side chain moieties. By using high-pressure NMR experiments, we clearly detected structural fluctuations in core domain of MSG which could not be observed in conventional NMR experiments.
Unfortunately, we could not do the quantitative analysis for structural dynamics of MSG and GroEL-GroES due to the lower sensitivity of exchange signals. We will continue to improve and develop the SAIL-NMR methods to solve these problems.

Free Research Field

構造生物学