Transient protein–protein interactions between cis-acting acyltransferase (AT) and acyl carrier protein (ACP) domains are critical for the catalysis and processivity of modular polyketide synthases (mPKSs), but are challenging for structural characterization due to the intrinsically weak binding affinity. Here, a stable complex of cis-acting AT and ACP domains from the ninth module of the salinomycin mPKS was obtained using a maleimide cross-linker and the structure of the complex was determined at 2.6 Å resolution. The crystal structure shows that the AT in combination with the ketosynthase (KS)-to-AT linker forms a C-shaped architecture to embrace the ACP. The large hydrolase subdomain of the AT serves as a major binding platform for the ACP, while the small ferredoxin-like subdomain of the AT and the KS-to-AT linker cooperate with each other to constrain binding of the ACP. The importance of interface residues in cis-acting AT–ACP interactions was confirmed by mutagenesis assays. The interaction mode observed in the cis-acting AT–ACP complex is completely different from those observed in trans-acting AT–ACP complexes, where the ACP primarily contacts the small domain of the AT. The complex structure provides detailed mechanistic insights into AT–ACP recognition in cis-AT mPKSs.
Celastrol has been identified as a potential candidate for anticancer drug development. In this study, 28 novel celastrol derivatives with C-6 sulfhydryl substitution and 20-substitution were designed and synthesized, and their antiproliferative activity against human cancer cells and non-malignant human cells was evaluated, with cisplatin and celastrol being used as controls. The results showed that most of the derivatives had enhanced in vitro anticancer activity compared to the parent compound celastrol. Specifically, derivative 2f demonstrated the most potent inhibitory potential and selectivity against HOS with an IC 50 value of 0.82 μM.Our study provides new insights into the structure-activity relationship of celastrol and suggests that compound 2f may be a promising drug candidate for the treatment of osteosarcoma.
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