Development of a simple selection method for interacting molecules using cell surface display.

2022 Fiscal Year Final Research Report

• Project/Area Number: 20K05889
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Review Section: Basic Section 38050:Food sciences-related
• Research Institution: Gifu University
• Principal Investigator: Ohno Satoshi 岐阜大学, 工学部, 准教授 (10345796)
• Project Period (FY): 2020-04-01 – 2023-03-31
• Keywords: 細胞表層工学

Outline of Final Research Achievements

In this study, we focused on the transpeptidase activity of Sortase for protein display to the cell surface layer of Brevibacillus sp. First, we measured the transpeptidase activity of Sortase using substrate peptides with various sorting signal motif sequences, and then we attempted to display a protein fused with HiBiT peptide to the cell surface layer. The amount of HiBiT peptide display on the cell surface was measured using LgBiT, and the signal intensity was stronger when the P22 secretory signal sequence was used. This result suggests that the target protein is displayed on the cell surface. Therefore, we mixed the fluorescent protein fusion LgBiT with cells and attempted to separate them by flow cytometry. Fluorescent protein-bound cells were detected, suggesting that it is possible to sort cells displaying the protein using interacting molecules.

Free Research Field

タンパク質工学

Academic Significance and Societal Importance of the Research Achievements

本研究では、ブレビバチルス菌の細胞表面に機能性分子を提示できることを示した。期間内に、アレルギーコンポーネントと相互作用する分子の取得までには至らなかったが、その基礎が構築できたことから、今後、食品検査等への応用が期待できる。また、選別された分子の調製において、ブレビバチルス菌は培養が容易なグラム陽性菌で、エンドトキシン活性を持たず、糖鎖などの翻訳後修飾もないことから、安全、安価で簡便に、高品質な分子の提供が期待できる。