The emergence and rapid spread of methicillin‐resistant Staphylococcus aureus (MRSA) raise a critical need for alternative therapeutic options. New antibacterial drugs and targets are required to combat MRSA‐associated infections. Based on this study, celastrol, a natural product from the roots of Tripterygium wilfordii Hook. f., effectively combats MRSA in vitro and in vivo. Multi‐omics analysis suggests that the molecular mechanism of action of celastrol may be related to Δ¹‐pyrroline‐5‐carboxylate dehydrogenase (P5CDH). By comparing the properties of wild‐type and rocA‐deficient MRSA strains, it is demonstrated that P5CDH, the second enzyme of the proline catabolism pathway, is a tentative new target for antibacterial agents. Using molecular docking, bio‐layer interferometry, and enzyme activity assays, it is confirmed that celastrol can affect the function of P5CDH. Furthermore, it is found through site‐directed protein mutagenesis that the Lys205 and Glu208 residues are key for celastrol binding to P5CDH. Finally, mechanistic studies show that celastrol induces oxidative stress and inhibits DNA synthesis by binding to P5CDH. The findings of this study indicate that celastrol is a promising lead compound and validate P5CDH as a potential target for the development of novel drugs against MRSA.

Alpha-Pharm3D, a multifunctional deep learning method, enables bioactivity predictions, elucidates PH4 fingerprints, and facilitates PH4-potential drug screening by explicitly incorporating geometric constraints.

The glucagon-like peptide-1 receptor (GLP-1R), which belongs to the class B1 G protein-coupled receptor (GPCR) family, is an important target for treatment of metabolic disorders, including type 2 diabetes and obesity. The growing interest in GLP-1R-based therapies is driven by the development of various functional agonists as well as the huge commercial market. Thus, understanding the structural details of ligand-induced signaling are important for developing improved GLP-1R drugs. Here, we investigated the conformational dynamics of the receptor in complex with a selection of prototypical functional agonists, including CHU-128 (small molecule-biased), danuglipron (small molecule balanced), and Peptide 19 (peptide balanced), which exhibit unique, distinct binding modes and induced helix packing. Furthermore, our all-atom molecular dynamics (MD) simulations revealed atomic feature how different those ligands led to signaling pathway preference. Our findings offer valuable insights into the mechanistic principle of GLP-1R activation, which are helpful for the rational design of next-generation GLP-1R drug molecules.