Covalent Modification of Proteins by Osthole Reactive Metabolites using Proteomic Approaches

Yue Zhuo; Huiling Chen; Chenchen Liu; Yida Zhang; Jiansong Fang; Meng Li; Zhendong Wang; Qiyao Jiang; Liangwen Yu; Huafeng Pan; Qi Wang

doi:10.2174/1389200224666230727123006

Covalent Modification of Proteins by Osthole Reactive Metabolites using Proteomic Approaches

Curr Drug Metab. 2023;24(8):611-620. doi: 10.2174/1389200224666230727123006.

Authors

Yue Zhuo¹, Huiling Chen¹, Chenchen Liu¹, Yida Zhang², Jiansong Fang¹, Meng Li³, Zhendong Wang¹, Qiyao Jiang¹, Liangwen Yu⁴, Huafeng Pan¹, Qi Wang¹

Affiliations

¹ Science and Technology Innovation Center, Guangzhou University of Chinese Medicine, Guangzhou, China.
² Department of Allergy and Clinical Immunology, National Center for Respiratory Medicine, National Clinical Research Center for Respiratory Disease, State Key Laboratory of Respiratory Disease, Guangzhou Institute of Respiratory Health, The First Affiliated Hospital of Guangzhou Medical University, Guangzhou, China.
³ Department of Pediatric Surgery, Guangzhou Institute of Pediatrics, Guangdong Provincial Key Laboratory of Research in Structural Birth Defect Disease, Guangzhou Women and Children's Medical Center, Guangzhou Medical University, Guangzhou, China.
⁴ Guangzhou University of Chinese Medicine Guangzhou University of Chinese Medicine Guangzhou China.

PMID: 37519003
DOI: 10.2174/1389200224666230727123006

Abstract

Background: Osthole (OST) is a bioactive natural coumarin derived from the plant Cnidium monnieri (L.) Cusson fruit (She Chuang Zi), which has various pharmacological and biological activities. OST contains an α,β- unsaturated lactone, which is an electrophilic group that tends to be metabolized into reactive metabolites (RMs). Then, RMs are able to covalently modify nucleophilic amino acid (AA) residues of target proteins. However, few researchers considered the contribution of the covalent modification induced by OST or its metabolites.

Objective: This study aims to investigate the metabolic profile and the metabolites-protein modification of OST.

Methods: The metabolites of OST were qualitatively identified using UHPLC-Q-TOF-MS. The RMs modification patterns and potentially modified AA residues were confirmed by UHPLC-Q-TOF-MS using rat liver microsomes (RLMs) and model AAs. Finally, the modified peptides derived from high-abundance microsomal peptides were separated via nano-LC-Orbitrap-MS, and then RM-modified proteins were identified using a proteome discoverer.

Results: In the presence of RLMs, OST could rapidly be metabolized within 1 h and hardly identified at 4 h. We detected 10 OST metabolites, 13 OST metabolites-NAC (N-acetyl cysteine) adducts, 3 NAL (N-acetyl lysine) adducts, and 11 GSH (glutathione) adducts. Furthermore, 16 RM-modified protein targets were identified, many of which are included in the essential biological processes of OST's anti-Alzheimer's disease (AD) and anti-tumor.

Conclusion: This study provides a novel perspective on the molecular mechanism of OST's pharmacological activities, as well as identifies potential targets for further development and application of OST and other Natural products (NPs).

Keywords: Covalent drugs; adducts; osthole; proteomics; reactive metabolites; traditional chinese medicine.