Perry Research Group

Research

Complex Coacervates as Biomimetic Materials

On Chip Laue

General schematic of molecular architectures for the formation of coacervates and structured coacervate domains. Specific molecular interactions (i.e., charge, hydrophobicity, hydrogen bonding) can be incorporated through the addition of different amino acid residues or other functionalities (not shown).

Living cells have evolved sophisticated inter- and intracellular organization, compartmentalization, transport, and signaling that are very challenging to reproduce synthetically. The functions of proteins and other macromolecules are dependent on both their individual structures, as well as the properties of the surrounding medium (i.e., viscosity, crowding, and other bulk phenomena, as well as direct molecular interactions). However, much of our understanding of the function and stability of biomolecules has been obtained in the absence of such effects, and has adversely affected the development of stabilized biomolecules for applications such as sensors, catalysis, and biomedicine. For instance, the storage and transportation of biomolecule-based vaccines requires refrigeration to maintain efficacy, while nature is able to stabilize proteins at physiological temperatures and higher.

One of the major challenges associated with the study of biological macromolecules is relating data obtained in the lab to the natural stability and activity of molecules in the crowded and confined intracellular environment. Functional enzyme studies, for instance, are typically performed in a dilute solution to minimize nonspecific interactions between molecules, while the actual in vivo environment contains ~20-40% protein by mass. Although the concentration of any single protein species may not be high, the total protein concentration can cause significant increases in the free energy of the solution compared to the dilute case. This increase in free energy is typically associated with decreases in entropy due to crowding. However, enthalpic effects related to chemical interactions with the environment may also be significant, but are far less studied.

Both theory and experiments have described the effects of crowding and confinement on the stability, folding, and activity of enzymes. Smaller, more compact states will be stabilized, consequently favoring or disfavoring subsequent reaction steps. Crowding also plays a significant role in driving the association of larger macromolecular assemblies and disfavoring denaturation. For instance, crowding increases the renaturation rate of DNA by 1 – 2 orders of magnitude, drives the association of 70S ribosomes from 30S and 50S particles, favors actin filament growth, and has been used to explain the need for molecular chaperones. Recently, confinement was also shown to play a role in stabilizing vaccines and antibiotics for long-term storage.

The current understanding of the specific molecular effects of crowding agents, both synthetic and natural, remains limited. This represents a major challenge in understanding and addressing the role of the environment surrounding a particular biomolecule. Commonly used polymers such as polyethylene glycol (PEG), dextran, and Ficoll, and related aqueous two-phase system (ATPS) strategies are unable to recapitulate the chemical composition of the cell interior. Unfortunately, using proteins directly as crowding agents, either in purified form or from cellular extracts, can result in unintended interactions. Thus, to develop an effective crowding medium, it is critical to identify and understand the possible interactions between the encapsulated biomolecule and the background matrix.

Specific molecular interactions such as electrostatics, hydrogen bonding, van der Waals forces, and preferential solvation result in enthalpic contributions to protein stability and/or activity. While individual effects from these interactions may be small, the synergy from a large number of combined effects can become significant, particularly given the degree of crowding present within intracellular spaces. Analysis and control of variations in biomolecule stability and activity as a function of the composition of the surrounding medium will enable identification of hierarchies of molecular interactions so that the complexity of the environment can be tailored to fit the needs of a particular experiment or application.

The most common model environments used for functional studies of biomolecules utilize polymers in solution to mimic crowding, and compartmentalization created by self-assembled vesicles or polymersomes. Creating a platform that combines both of these effects has remained a challenge for a variety of reasons including the impermeability of many lipid or polymer membranes to solutes, and the harsh solvent conditions associated with the assembly of many synthetic polymer systems. We are investigating a strategy utilizing complex coacervation to drive the membraneless sequestration and organization of biomolecules and biomolecular networks within a biomimetic intracellular environment where complex molecular interactions can be built in by design.

Complex coacervation is a phase separation phenomenon resulting from the electrostatic complexation of oppositely-charged polyelectrolytes. The resultant coacervate is a dense, polymer-rich liquid retaining both water and salt, and are common in everyday life applications such as processed food, cosmetics, Kindle® displays, and the natural adhesives used by sessile marine animals. Molecular design can be coupled with coacervation, linking polyelectrolyte domains to neutral polymer blocks to stabilize microphase separation of coacervate domains (see Figure) into structures associated with traditional block copolymers (i.e., micelles, rods, bicontinuous). Both hydrophobic and hydrophilic molecules can be loaded into these coacervate domains, facilitating drug delivery and diagnostics. Emerging experience has shown that polypeptide-based coacervation produces a dense amino acid-rich phase that closely mimics protein concentrations within cells and is an effective medium for protein encapsulation and stabilization.

Funding

Collaborations

References

  1. W.C. Blocher, S.L. Perry, "Biomimetic Complex Coacervate-Based Materials for Biomedicine," WIREs Nanomedicine and Nanobiotechnology, (2017), 9(4), e1442. [PDF].
  2. X. Meng, S.L. Perry, J.D. Schiffman, "Complex Coacervation: Chemically Stable Fibers Electrospun from Aqueous Polyelectrolyte Solutions," ACS Macro Letters, (2017), 6, 505-511. [PDF]
  3. Highlighted in: Data from University of Massachusetts Advance Knowledge in Tissue Engineering, Biotech Week, June 28th, 2017.

  4. M. Radhakrishna, K. Basu, Y. Liu, R. Shamsi, S.L. Perry, C.E. Sing, "Molecular Connectivity and Correlation Effects on Polymer Coacervation," Macromolecules, (2017) 50(7), 3030–3037. [PDF]
  5. Y. Liu, H.H. Winter, S.L. Perry, "Linear Viscoelasticity of Complex Coacervates," Advances in Colloid and Interface Science, (2017) 239, 46-60. [PDF]
  6. D. Priftis, L. Leon, Z. Song, S.L. Perry, K.O. Margossian, A. Tropnikova, J. Cheng, M. Tirrell, "Self-Assembly of α-Helical Polypeptides Driven by Complex Coacervation," Angewandte Chemie International Edition, (2015) 54, 11128-11132. [PDF]
  7. S.L. Perry, C.E. Sing, "PRISM-Based Theory of Complex Coacervation: Excluded Volume versus Chain Correlation," Macromolecules, 48, 5040-5053 (2015). [PDF]
  8. K.Q. Hoffmann, S.L. Perry, L. Leon, D. Priftis, M. Tirrell, J.J. de Pablo, "A Molecular View of the Role of Chirality in Charge-driven Polypeptide Complexation," Soft Matter, 11, 1525-1538 (2015). [PDF]
  9. S.L. Perry, L. Leon, K.Q. Hoffmann, M.J. Kade, D. Priftis, K.A. Black, D. Wong, R.A. Klein, C.F. Pierce, K.O. Margossian, J.K. Whitmer, J. Qin, J.J. de Pablo, and M. Tirrell, "Chirality Selected Phase Behavior in Ionic Polypeptide Complexes," Nature Communications, 6, 6052 (2015). [PDF]
  10. D.V. Krogstad, S.H. Choi, N.A. Lynd, D.J. Andrus, S.L. Perry, J.D. Gopez, C.J. Hawker, E.J. Kramer, M. Tirrell, "Small Angle Neutron Scattering Study of Complex Coacervate Micelles and Hydrogels Formed from Ionic Diblock and Triblock Copolymers," Journal of Physical Chemistry B, 118, 13011-13018 (2014). [PDF]
  11. K.A. Black, D. Priftis, S.L. Perry, J. Yip, W.Y. Byun, M. Tirrell, "Protein Encapsulation via Polypeptide Complex Coacervation," ACS Macro Letters, 3, 1088-1091 (2014). [PDF]
  12. Highlighted on the C&EN Biological and Materials News SCENE.

    Highlighted in: "Charged Polymers Package Proteins," Chemical & Engineering News, 92(45), 27 (2014).

  13. S.L. Perry, Y. Li, D. Priftis, L. Leon, M. Tirrell, "The Effect of Salt on the Complex Coacervation of Vinyl Polyelectrolytes," Polymers, 6, 1756-1772 (2014). [PDF]
  14. J. Qin, D. Priftis, R. Farina, S.L. Perry, L. Leon, J. Whitmer, K. Hoffmann, M. Tirrell, J.J. de Pablo, "Interfacial Tension of Polyelectrolyte Complex Coacervate Phases,"ACS Macro Letters, 3, 565-568 (2014). [PDF]
  15. D. Priftis, X. Xia, K.O. Margossian, S.L. Perry, L. Leon, J. Qin, J.J. de Pablo, M. Tirrell, "Ternary, Tunable Polyelectrolyte Complex Fluids Driven by Complex Coacervation," Macromolecules, 47(9), 3076-3085 (2014). [PDF]