The growing threat of multidrug-resistant bacterial infections demands innovative therapeutic strategies that transcend traditional small-molecule antibiotic development (Rathored J, Budhbaware T., 2025). Lactoferrin (LF), a multifunctional iron-binding glycoprotein, offers a promising biomolecular scaffold owing to its intrinsic antimicrobial and immunomodulatory activities (Legrand D, 2016). Here, we integrate structural biology, protein engineering, and nanomedicine to develop a novel protein-based drug delivery platform derived from the lactoferrin C-lobe.
To establish a stable and well-defined building block, we solved the high-resolution crystal structure of the chymotrypsin-cleaved C-lobe (CLF). Our structural analysis confirms that the isolated domain retains its native tertiary fold and conformational integrity post-cleavage, providing a validated atomic-level framework for subsequent engineering and rational nanocarrier design.
Leveraging this structurally verified scaffold, we successfully transformed both full-length LF and the CLF into stable nanoparticles using a simple, biocompatible sol-oil method with olive oil as a stabilizing agent (Kumar P, 2017). Both nanoparticle systems demonstrated high encapsulation efficiencies for the model antimicrobials moxifloxacin and curcumin, with curcumin exhibiting superior loading due to its hydrophobic character. Crucially, SDS-PAGE and long-term stability assays confirmed that both protein scaffolds maintained structural integrity throughout nanoparticle fabrication, drug loading, and after one year of storage, underscoring the robustness of our engineering approach.
When evaluated against clinically relevant bacterial pathogens, drug-loaded nanoparticles exhibited significantly enhanced antimicrobial potency. LF-curcumin nanoparticles achieved up to a 15-fold reduction in the minimum inhibitory concentration (MIC) against Staphylococcus aureus, while LF-moxifloxacin nanoparticles reduced the MIC against Acinetobacter baumannii by 2.5-fold. Intriguingly, while CLF nanoparticles displayed superior physicochemical properties, including smaller particle size and more favourable zeta potential, the intact LF nanoparticles consistently yielded greater antimicrobial synergy, suggesting an essential functional contribution from the N-lobe. Notably, CLF nanoparticles did not compromise antibiotic efficacy, validating their utility as alternative proteinaceous nanocarriers.
Collectively, this work establishes the structurally validated lactoferrin C-lobe as a stable, engineerable protein scaffold for antimicrobial delivery. By bridging atomic-resolution structural biology with applied nanotechnology, our findings provide a rational blueprint for the development of lactoferrin-derived nanomedicines to combat recalcitrant bacterial infections.
Reference
Rathored J, Budhbaware T. Integrative strategies against multidrug-resistant bacteria: Synthesizing novel antimicrobial frontiers for global health. Microb Pathog. 2025;208:108018. doi:10.1016/j.micpath.2025.108018
Legrand D. Overview of Lactoferrin as a Natural Immune Modulator. J Pediatr. 2016;173 Suppl:S10-S15. doi:10.1016/j.jpeds.2016.02.071
Kumar P, Lakshmi YS, Kondapi AK. An oral formulation of efavirenz-loaded lactoferrin nanoparticles with improved biodistribution and pharmacokinetic profile. HIV Med. 2017;18(7):452-462. doi:10.1111/hiv.12475