Study of the stabilizing effect of different solvents on the green fluorescent protein structure produced by Escherichia coli

• Grant number: 18/01858-2
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Start date: June 01, 2018
• End date: August 09, 2020
• Field of knowledge: Engineering - Chemical Engineering - Chemical Technology
• Principal Investigator: Valéria de Carvalho Santos Ebinuma
• Grantee: Carolina Falaschi Saponi
• Host Institution: Faculdade de Ciências Farmacêuticas (FCFAR). Universidade Estadual Paulista (UNESP). Campus de Araraquara. Araraquara , SP, Brazil
• Associated scholarship(s): 18/20833-0 - Entendendo as interações moleculares e a estabilidade da proteína verde fluorescente (GFP) em líquidos iônicos sob condições de estresse, BE.EP.IC

Abstract

The Green Fluorescent Protein (GFP) has intense and natural fluorescence, being easily quantifiable and having excellent application as biosensor and biomarker. However, the manifestation of its fluorescence is dependent on maintaining the integrity of its protein structure, which is very sensitive to different stress conditions, such as high temperatures, extreme pH values or the presence of denaturing substances. GFP instability under various conditions, coupled with its high cost of production, somehow restricts its use to small-scale, generally academic projects. Thus, the study of solvents that allow the protection of GFP structure under stress conditions is essential to allow its use on a large scale. Considering this information, the main objective of the project is to evaluate the stability of the recombinant GFP produced by Escherichia coli in different solvents and under stress conditions, in order to identify compounds with the capacity to stabilize proteins and/or that lead to the increase/suppression of fluorescence (for possible application in the development of biosensors). For this purpose, fluorescence analysis techniques (3D fluorescence spectroscopy and fluorescence intensity analysis) will be used and, to obtain additional knowledge about the solvent-protein interactions, circular dichroism and/or Attenuated Total Reflectance Fourier Transform Infrared(ATR-FTIR) will also be used. The solvents to be used will be polymers, deep eutectic solvents (DESs) and different families of ionic liquids (LIs).

Scientific publications

(References retrieved automatically from Web of Science and SciELO through information on FAPESP grants and their corresponding numbers as mentioned in the publications by the authors)

DOS SANTOS, NATHALIA VIEIRA; SAPONI, CAROLINA FALASCHI; GREAVES, TAMAR LOUISE; BRANDAO PEREIRA, JORGE FERNANDO. Revealing a new fluorescence peak of the enhanced green fluorescent protein using three-dimensional fluorescence spectroscopy. RSC ADVANCES, v. 9, n. 40, p. 22853-22858, AUG 1 2019. (14/16424-7, 18/50009-8, 18/06576-5, 18/01858-2, 14/19793-3, 16/07529-5)

DOS SANTOS, NATHALIA VIEIRA; SAPONI, CAROLINA FALASCHI; RYAN, TIMOTHY M.; PRIMO, FERNANDO L.; GREAVES, TAMAR L.; PEREIRA, JORGE F. B.. Reversible and irreversible fluorescence activity of the Enhanced Green Fluorescent Protein in pH: Insights for the development of pH-biosensors. International Journal of Biological Macromolecules, v. 164, p. 3474-3484, DEC 1 2020. (18/20833-0, 14/16424-7, 14/19793-3, 18/06576-5, 16/07529-5, 18/50009-8, 18/01858-2)

V. VERISSIMO, NATHALIA; SAPONI, CAROLINA F.; RYAN, TIMOTHY M.; GREAVES, TAMAR L.; PEREIRA, JORGE F. B.. Imidazolium-based ionic liquids as additives to preserve the Enhanced Green Fluorescent Protein fluorescent activity. GREEN CHEMICAL ENGINEERING, v. 2, n. 4, p. 11-pg., 2021-12-01. (18/06576-5, 18/25511-1, 14/16424-7, 20/14144-8, 18/50009-8, 14/19793-3, 18/01858-2, 18/20833-0, 16/07529-5)