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 Final Research Report

• Project/Area Number: 19K16348
• Research Category: Grant-in-Aid for Early-Career Scientists
• Allocation Type: Multi-year Fund
• Review Section: Basic Section 47030:Pharmaceutical hygiene and biochemistry-related
• 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 / Keap1 / GSTP1 / drug-induced toxicity

Outline of Final Research Achievements

GSTP1 catalyzes glutathionylation of proteins to alter their functions. Under normal condition, Keap1 protein interacts with Nrf2 protein, which induces detoxification-related genes, to interfere the translocation into nucleus. Upon oxidative stress, Keap1 is suggested to be glutathionylated, which may affect the interaction with Nrf2 protein. Several experiments were conducted to understand molecular mechanisms, and the following results were obtained. 1. Purified human GSTP1, Keap1, and Nrf2 were successfully obtained from E.coli heterologous expression system. 2. HS-AFM imaging conditions for these proteins were established and optimized. This is the first observation of these dynamic structure at single molecule level. 3. Clear oxidants-dependent glutathionylation of Keap1 were observed, which also indicating an oxidative status-dependency of Keap1 molecular structure. With application of this method, glutathionylated protein concentrations were determined from in vitro samples.

Free Research Field

薬系衛生および生物化学関連

Academic Significance and Societal Importance of the Research Achievements

The elucidation of dynamic structures of Keap1 and Nrf2 is helpful for the understanding of their physiological roles and regulatory roles in oxidative stress status. The regulation of Keap1 glutathionylation on interaction with Nrf2 provides new insights into mechanisms of drug-induced toxicity.