Characterization of human glutathione S-transferase P1-1-catalyzed glutathionylation of proteins: as a clue to understand the inter-individual difference in drug-induced toxicity

2020 Fiscal Year Annual Research Report

• Project/Area Number: 19K16348
• Research Institution: Kanazawa University
• Principal Investigator: ZHANG YONGJIE 金沢大学, ナノ生命科学研究所, 協力研究員 (80836180)
• Project Period (FY): 2019-04-01 – 2021-03-31
• Keywords: glutathionylation / Nrf2 / Keap1 / HS-AFM

Outline of Annual Research Achievements

We have established the heterologous expression systems for human nuclear factor erythroid 2-related factor 2 (Nrf2) to visualize the interaction with Kelch-like ECH-associated protein 1 (Keap1) using high-speed atomic force microscopy (HS-AFM). And then, His-tagged Nrf2 protein was tried to be purified with a series of column chromatographies, but Nrf2 protein with high purity to observe with HS-AFM could not be obtained. However, we tried to visualize Nrf2 itself with HS-AFM, resulting that Nrf2 protein could be observed with the shape (large two domains at N-terminus and one small domain at C-terminus) predicted using Predictor of Natural Disordered Region site. Thus, we firstly succeeded to visualize Nrf2 protein with HS-AFM. However, due to the impurity of Nrf2 protein, it was difficult to conduct the subsequent observation. Particularly, the effect of glutathionylation of Keap1 on the interaction with Nrf2 was of interest. To more clearly observe Nrf2 protein and the interaction with Keap1 protein, we should obtain Nrf2 protein with high purity with an improved methods.