Bridging the gap between lactoferrin and V-ATPase through a multi-stage computational approach

Lactoferrin (Lf), a bioactive milk protein, exhibits strong anticancer and antifungal activities [1,2]. The search for Lf targets and mechanisms of action is of utmost importance to enhance its effective applications. A common feature among Lf-treated cancer and fungal cells is the inhibition of a proton pump essential for pH homeostasis called V-ATPase. Lf-driven V-ATPase inhibition leads to cytosolic acidification, ultimately causing cell death of cancer and fungal cells [2–4]. Given that a detailed elucidation of how Lf and V-ATPase interact is still missing, in this work we aimed to fill this gap by employing a multi-level computational approach. Molecular dynamics (MD) simulations of both proteins were performed to obtain a robust sampling of their conformational landscape, followed by clustering and protein-protein docking. Subsequently, MD simulations of the docked complexes and free binding energy calculations were carried out to evaluate the dynamic binding process and built the final ranking. This computational pipeline unraveled a putative mechanism by which Lf inhibits V-ATPase and identified key binding residues that will certainly aid in the rational design of follow-up experimental studies, bridging in this way computational and experimental biochemistry.

1. Acosta-Zaldívar M, Andrés MT, Rego A, Pereira CS, Fierro JF, Côrte-Real M. Human lactoferrin triggers a mitochondrial- and caspase-dependent regulated cell death in Saccharomyces cerevisiae. Apoptosis. 2016;21: 163–173. doi:10.1007/s10495-015-1199-9
2. Pereira CS, Guedes JP, Gonçalves M, Loureiro L, Castro L, Gerós H, Rodrigues LR, Côrte-Real M. Lactoferrin selectively triggers apoptosis in highly metastatic breast cancer cells through inhibition of plasmalemmal V-H+-ATPase. Oncotarget. 2016;7: 62144–62158. doi:10.18632/oncotarget.11394
3. Guedes JP, Pereira CS, Rodrigues LR, Côrte-Real M. Bovine milk lactoferrin selectively kills highly metastatic prostate cancer PC-3 and osteosarcoma MG-63 cells in vitro. Front Oncol. 2018;8: 1–12. doi:10.3389/fonc.2018.00200

4. Pereira CS, Andrés MT, Chaves SR, Fierro JF, Gerós H, Manon S, Rodrigues LR, Côrte-Real M. Lactoferrin perturbs lipid rafts and requires integrity of Pma1p-lipid rafts association to exert its antifungal activity against Saccharomyces cerevisiae. Int J Biol Macromol. 2021;171: 343–357. doi:10.1016/j.ijbiomac.2020.12.224