Overcoming the problems in CryoEM analysis using sugar chain and its application to the innate immune receptor, TLR

2022 Fiscal Year Final Research Report

• Project/Area Number: 21K19328
• Research Category: Grant-in-Aid for Challenging Research (Exploratory)
• Allocation Type: Multi-year Fund
• Review Section: Medium-sized Section 47:Pharmaceutical sciences and related fields
• Research Institution: The University of Tokyo
• Principal Investigator: Shimizu Toshiyuki 東京大学, 大学院薬学系研究科(薬学部), 教授 (30273858)
• Project Period (FY): 2021-07-09 – 2023-03-31
• Keywords: クライオ電子顕微鏡 / Orientation bias / 糖鎖

Outline of Final Research Achievements

We tried to solve the problems in CryoEM analysis (aggregation, orientation bias etc) by the chemical modification (PEGylation). PEGylation effectivly work in some proteins, allowing the structural analysis. TLR7 is a glycoprotein and we prepared the TLR7 trimmed by EndH, and no trimmed TLR7. However, orientation bias in CryoEM analysis was not improved in both cases. In addition, introduction of glyco-sites sometimes affect the expression level of the protein.

Free Research Field

構造生物学

Academic Significance and Societal Importance of the Research Achievements

PEG化などの化学修飾は容易に試みることができる方法であり、クライオ電子顕微鏡解析で問題点がある場合は試す価値のある方法である。一方糖鎖導入は手間のかかる方法でありOrientation biasを解決するまでにはいたらないが、タンパク質の物性を変えることも期待できるので試行する価値はあると考えられる。