Polymeric crowding agents improve passive biomacromolecule encapsulation in lipid vesicles

Area of Science:

• Biotechnology and Nanotechnology
• Materials Science
• Physical Chemistry

Background:

• Encapsulating large biomacromolecules (e.g., proteins) into lipid vesicles is challenging due to low passive trapping efficiency.
• High molecular weight solutes often result in significantly lower internal vesicle concentrations compared to external solutions.

Purpose of the Study:

• To investigate the impact of macromolecular crowding on the passive encapsulation of biological macromolecules.
• To determine if crowding agents can enhance the encapsulation efficiency of large solutes in giant lipid vesicles (GVs) and large unilamellar vesicles (LUVs).

Main Methods:

• Investigated encapsulation of fluorescently labeled biomacromolecules (52 kDa to 660 kDa) during vesicle formation.
• Utilized poly(ethylene glycol) (PEG) and dextran as crowding agents (3 wt%).
• Compared encapsulation efficiency in the presence and absence of crowding agents for GVs and LUVs.

Main Results:

• Lower molecular weight biomacromolecules (hydrodynamic radius < 10 nm) showed encapsulation efficiency equal to external concentrations.
• Larger biomacromolecules (hydrodynamic radius >= 10 nm) exhibited markedly reduced internal concentrations.
• Addition of PEG or dextran significantly improved encapsulation of larger biomacromolecules to levels matching smaller ones.

Conclusions:

• Macromolecular crowding agents (PEG, dextran) act as volume excluders, effectively increasing biomacromolecule encapsulation.
• This strategy provides a simple and general method to enhance the loading of poorly encapsulated macromolecular solutes into both GVs and LUVs.
• Achieved internal solute concentrations can match external solution concentrations, overcoming previous limitations.