Polyelectrolyte micelles: characterization, properties and applications for enzymes encapsulation

Abstract

Inside cells, the functions performed by proteins and other biomolecules depend not only on their intrinsic characteristics, but also on properties of the medium, such as viscosity, water content and ion concentration. Many biological systems present the process of associative phase separation between polyelectrolytes, forming structures called membraneless organelles. This process can be mimicked by the complexation between oppositely charged polyelectrolytes, which forms a biphasic system, with a rich phase in the colloidal components (the coacervate). However, if the coacervation process involves a charged-neutral diblock copolymer, the coacervate is confined to colloidal dimensions, in a core-shell structure, called complex coacervate core micelle, or C3M. The interior of C3M structures can act as a compartment of biomolecules, thus it mimics the so-called membraneless organelles. Several aspects related to the compartmentalization of biomolecules are of fundamental importance, such as the stability and structure of the biomolecule-C3M complex. In this sense, this project seeks to investigate both these fundamental issues, preparing aggregates containing enzymes compartmentalized in the core. Possible synergistic or antagonistic effects on the enzymatic activity will be studied when the enzyme is encapsulated inside the coacervate. It will also be prepared a recoverable enzymatic nanoreactor. For this, a specific enzyme-C3M complex will be investigated, in which the neutral hydrophilic block of the copolymer is thermo-responsive. Thus, after the enzymatic catalysis, the coacervate-enzyme can be separated from the reaction medium by heating the system, resulting in the formation of a macrophase. With the subsequent cooling, the coacervate structure returns to its original form, allowing for a new reaction cycle. Techniques such as light scattering, x-ray scattering at low angle, isothermal titration calorimetry and nuclear magnetic resonance spectroscopy will be used to monitor the formation of the C3M protein structure. The enzymatic cycles will be studied by evaluating the kinetics and reaction yield, monitoring the concentrations of substrate and product. (AU)