Interfacial Assembly of Bacterial Microcompartment Shell Proteins in Aqueous Multiphase Systems
A A Dharani T Abeysinghe1, Eric J Young2,3, Andrew T Rowland1
1Department of Chemistry, Pennsylvania State University, State College, PA, 16801, USA.
Small (Weinheim an Der Bergstrasse, Germany)
|December 1, 2023
Summary
Researchers combined bacterial microcompartment (BMC) shell proteins with liquid-liquid phase separation (LLPS) to create novel compartments. These protein-coated droplets offer a new strategy for synthetic cells, organizing biomimetic functions with cargo like enzymes or RNA.
Area of Science:
- Biochemistry
- Synthetic Biology
- Materials Science
Background:
- Compartmentalization is essential for life, achieved through various structures like lipid membranes, protein shells, or biopolymer phase separation.
- Bacterial microcompartment (BMC) shell proteins are known for self-assembly, forming protein-based compartments within bacteria.
- Liquid-liquid phase separation (LLPS) is a biophysical process creating membrane-less compartments through spontaneous de-mixing of cellular components.
Purpose of the Study:
- To develop novel forms of compartmentalization by integrating BMC shell proteins with LLPS.
- To investigate the assembly behavior of BMC proteins at liquid-liquid interfaces in different phase-separated systems.
- To explore the potential of these engineered compartments for organizing biomimetic functionality within synthetic cells.
Main Methods:
- Utilized poly(ethyleneglycol)/dextran aqueous two-phase systems and polylysine/polyaspartate complex coacervate systems to create liquid-liquid interfaces.
- Assembled bacterial microcompartment (BMC) shell proteins at the interfaces of these phase-separated systems.
- Investigated the influence of polyelectrolyte ratios and protein concentrations on interfacial assembly and competition with aggregation.
- Developed a three-phase system by encapsulating coacervate droplets within dextran-rich droplets for tunable protein localization.
Main Results:
- BMC shell proteins successfully assembled at the liquid-liquid interfaces of both PEG/dextran and coacervate systems.
- Interfacial assembly in coacervate systems was sensitive to the ratio of cationic to anionic polypeptides, indicating electrostatic control.
- Protein concentration and polycation availability influenced the competition between interfacial assembly and aggregation.
- A three-phase system demonstrated tunable interfacial localization of BMC proteins by adjusting the polyelectrolyte charge ratio.
Conclusions:
- BMC shell proteins can be directed to assemble at liquid-liquid interfaces, creating novel protein-coated droplets.
- These engineered droplets, formed via LLPS and protein self-assembly, represent a new strategy for synthetic cell design.
- The ability to encapsulate cargo like enzymes or RNA within these compartments enables the organization of biomimetic functions.
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