Tito, Gabriella (2025) Protein/DNA metalation: a combined experimental/computational approach. [Tesi di dottorato]

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Abstract

Understanding how metallodrugs interact with biological macromolecules is essential for elucidating their mechanisms of action and for the rational design of new therapeutic agents. Bis(acetylacetonato)oxidovanadium(IV), [V(IV)O(acac)₂], is one of the most extensively investigated vanadium-based drugs due to its ability to decrease blood glucose levels and enhance the kinase activity of the insulin receptor. Dirhodium tetraacetate is a bimetallic complex with anticancer potential. Protein binding can modulate their bioactivity by affecting blood transport and mechanism of action. For this reason, it is important to study the interaction of these complexes with proteins. Thus, the structures of the adducts that [V(IV)O(acac)₂] forms with the model protein hen egg white lysozyme (HEWL) and the physiologically relevant protein human H-chain ferritin (HhFt) were studied by high-resolution X-ray crystallography and using spectroscopic techniques and computational methods. The structures with HEWL were obtained at both 100 K and 37 °C. In all cases, the reaction of [V(IV)O(acac)₂] with the proteins leads to the formation of unexpected and unusual polyoxidovanadates (POV) clusters. The structure of dirhodium tetraacetate with HEWL was also solved at 37 °C and compared with that already reported and solved at 100K. Results indicate only subtle temperature-dependent variations in metal occupancy and local residue mobility can be observed. The interaction of metallodrugs with DNA establishes another key element of their biological activity. High-resolution structural studies on adducts formed with several anticancer platinum-based compounds (cisplatin, transplatin, arsenoplatin, carboplatin, oxaliplatin) and with dirhodium tetraacetate with B-DNA revealed defined different coordination sites. Platinum centres preferentially bind N7 atom of guanine, while rhodium centre binds N7 atom of adenine. A comprehensive analysis of metal–DNA adducts in the Protein Data Bank indicates that detailed structural information on nucleic acid–metallodrug interactions remains limited. To address this gap, a new molecular docking protocol was developed to predict covalent interactions between metallodrugs and DNA. Overall, these findings provide valuable structural insights into metal-biomolecule interactions and introduce a powerful computational tool to guide the design of next-generation DNA-targeting metallodrugs.

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