Mapping mRNA Localization and Internal Structure in Lipid Nanoparticles through Solid-State Dynamic Nuclear...

Area of Science:

• Biophysics
• Materials Science
• Nanotechnology

Background:

• Lipid nanoparticles (LNPs) are crucial for RNA vaccine delivery.
• The internal structure of LNPs, especially with mRNA, is not well understood.
• Understanding LNP structure is key for optimizing RNA delivery systems.

Purpose of the Study:

• To investigate the internal structure and component distribution within intact LNPs.
• To apply relayed dynamic nuclear polarization (DNP)-enhanced solid-state NMR for structural analysis.
• To provide a quantitative structural framework for designing improved RNA delivery systems.

Main Methods:

• Relayed dynamic nuclear polarization (DNP)-enhanced solid-state NMR spectroscopy.
• Optimization of cryoprotectant removal and polarizing matrix composition.
• Numerical simulations of spin diffusion and polarization transport.

Main Results:

• Direct observation of encapsulated mRNA within functional LNPs using solid-state NMR.
• Quantitative analysis revealed a concentric LNP architecture: PEG-lipids at the surface, helper lipids in intermediate layers, and mRNA in the core.
• A multi-layered core-shell model best explained the NMR data and simulations.

Conclusions:

• Relayed-DNP MAS NMR is a powerful technique for probing nanoscale organization in LNPs.
• LNPs exhibit a defined core-shell structure with stratified lipid layers surrounding the mRNA core.
• This structural understanding can guide the rational design of more effective RNA delivery systems.