Study of lysozyme adsorption and desorption in layer-by-layer films by means of FRET

• Grant number: 19/06677-9
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Start date: June 01, 2019
• End date: May 31, 2020
• Field of knowledge: Physical Sciences and Mathematics - Chemistry - Organic Chemistry
• Principal Investigator: Luiz Henrique Catalani
• Grantee: Raphael Colonese Vlasman
• Host Institution: Instituto de Química (IQ). Universidade de São Paulo (USP). São Paulo , SP, Brazil
• Associated research grant: 18/13492-2 - Synthetic and natural scaffolds applied to regenerative medicine, AP.TEM

Abstract

This project aims to understand the adsorption and desorption processes of lysozyme (LYS) in Layer-by-Layer (LbL) self-assembly films of heparin (HEP; negative monolayer) and chitosan (CHI; positive monolayer) and built upon silicon wafers and PLLA films. For the adsorption study, both the protein and polyelectrolyte will be functionalized with Fluorescence Resonance Energy Transfer (FRET) donor-acceptor pair: LYS with fluorescein isothiocyanate (FITC), the acceptor probe, and HEP with dansyl chloride, the donor probe. After the development of efficient functionalization, multilayer films will be made by LbL self-assembly in both Silicon and PLLA films to follow the kinetics and mechanism of adsorption by changes in the fluorescence intensity. After, different conformations of the multilayers will be tested to look into the protein migration between films of polyelectrolytes. In this step, the heights of the functionalized HEP and LYS in the films will be changed among initial, intermediary and final layers. Lastly, the LYS release study will be done varying its position in the multilayer and so develop control over the release kinetics. The main goal is to verify the formation of coacervates between the proteins and the polyelectrolytes and how this event can influence the mechanism of adsorption and desorption of the proteins from the LbL system.