Comprehensive Analysis of the Multi-Target Binding Mechanism of Doxorubicin: Integrating Protein Microarray...

Understanding the mechanisms through which anticancer drugs interact with multiple protein targets is crucial for optimizing drug design and enhancing the efficacy of chemotherapy. This study focuses on doxorubicin, a broad-spectrum anticancer drug recognized for its multi-target mechanisms of action. We initially screened 363 doxorubicin-binding proteins using protein microarrays; of these, 166 proteins with known PDB (Protein Data Bank) structures were selected for molecular docking to evaluate their binding energies. The binding energy distribution and residue enrichment analyses revealed that doxorubicin preferentially binds to specific residues at its binding sites, including serine, glycine, arginine, glutamic acid, lysine, aspartic acid, and leucine. These residues stabilize doxorubicin binding through hydrogen bonds, hydrophobic interactions, and electrostatic interactions. In addition, RUVBL1 (RuvB-like AAA ATPase 1) exhibited the highest integrated score from the protein microarray and molecular docking analyses. Furthermore, PPI (protein-protein interaction) network analysis and centrality calculations identified key proteins with potential regulatory roles, with MAPK1 (mitogen-activated protein kinase 1) exhibiting the highest betweenness centrality in the PPI network. Finally, molecular dynamics simulations of the RUVBL1- and MAPK1-doxorubicin complexes were conducted to evaluate the binding mechanisms. The simulations revealed key binding residues, including Ile56, Lys59, Leu87, Pro296, and Ile326 in RUVBL1 and Asp88, Ile89, Pro93, Phe354, and Ala92 in MAPK1 that mediate stable interactions with doxorubicin. This study presents a comprehensive analytical approach for investigating the interactions between doxorubicin and multiple protein targets, providing a reference framework for understanding the molecular mechanisms of anticancer drugs and for future analyses of similar data sets.