Development of high-throughput and high-sensitivity 3D structure analysis method for proteins by mass spectrometry

2022 Fiscal Year Final Research Report

• Project/Area Number: 21K14652
• Research Category: Grant-in-Aid for Early-Career Scientists
• Allocation Type: Multi-year Fund
• Review Section: Basic Section 34020:Analytical chemistry-related
• Research Institution: Kyoto University
• Principal Investigator: Eisuke Kanao 京都大学, 薬学研究科, 助教 (40895166)
• Project Period (FY): 2021-04-01 – 2023-03-31
• Keywords: 分子インプリント法 / プロテオミクス / 構造解析

Outline of Final Research Achievements

We synthesized a hydrogel that effectively captured target proteins in aqueous conditions by using molecular imprinting technique. To enable selective fluorescence detection of the protein, we introduced a functional monomer that exhibits fluorescence upon hydrophobic interaction with the protein. Additionally, by incorporating a photoreactive monomer, perfluorophenyl azide, we developed a photoresponsive molecularly imprinted hydrogel, which enables selective capture of the protein and chemical bonding to its surface. By evaluating the enzyme-digested peptides of the captured protein within the hydrogel using LC-MS/MS, we successfully identified the surface primary structures of the protein.

Free Research Field

分析化学

Academic Significance and Societal Importance of the Research Achievements

本研究で提案した手法は，分子インプリント法を利用することで，LC-MS/MSによる一次構造解析を高次構造解析に応用するものであり，X線結晶解析をはじめとするタンパク質高次構造解析法とは感度と速度の面で一線を画すものである。そのため，生命科学的に重要な機能を担うタンパク質種や複合体をアミノ酸レベルで素早く簡便に予測し，分光学的手法での高次構造分析が必要なターゲットを絞り込むことができる。このように本研究は，既存の分光学的手法が抱える操作の煩雑さ・適用範囲の狭さを補完することが可能であり，ポストゲノム研究の中核を担う構造生物学分野の進展に大いに貢献できる。