rotic Ionic Liquid Cation Alkyl Chain Length Effect on Lysozyme Structur

Author(s)

Han, Qi ^[1] ; Broomhall, Hayden C. ^[1] ; Vieira Verissimo, Nathalia ^[2] ; Ryan, Timothy M. ^[3] ; Drummond, Calum J. ^[1] ; Pereira, Jorge F. B. ^[4] ; Greaves, Tamar L. ^[1]

Affiliation

^[1] RMIT Univ, Sch Sci, STEM Coll, 124 La Trobe St, Melbourne, Vic 3000 - Australia

^[2] Sao Paulo Univ USP, Sch Pharmaceut Sci, Ave Prof Lineu Prestes 580, Cidade Univ, BR-05508000 Sao Paulo - Brazil

^[3] Australian Nucl Sci & Technol Org, Australian Synchrotron, 800 Blackburn Rd, Clayton, Vic 3168 - Australia

^[4] Univ Coimbra, CIEPQPF, Dept Chem Engn, Rua Silvio Lima, Polo II Pinhal de Marrocos, P-3030790 Coimbra - Portugal

• Document type: Journal article
• Source: OLECULE; v. 27, n. 3 FEB 2022.
• Web of Science Citations: 0

Abstract

Solvents that stabilize protein structures can improve and expand their biochemical applications, particularly with the growing interest in biocatalytic-based processes. Aiming to select novel solvents for protein stabilization, we explored the effect of alkylammonium nitrate protic ionic liquids (PILs)-water mixtures with increasing cation alkyl chain length on lysozyme conformational stability. Four PILs were studied, that is, ethylammonium nitrate (EAN), butylammonium nitrate (BAN), hexylammonium nitrate (HAN), and octylammonium nitrate (OAN). The surface tension, viscosity, and density of PIL-water mixtures at low to high concentrations were firstly determined, which showed that an increasing cation alkyl chain length caused a decrease in the surface tension and density as well as an increase in viscosity for all PIL solutions. Small-angle X-ray scattering (SAXS) was used to investigate the liquid nanostructure of the PIL solutions, as well as the overall size, conformational flexibility and changes to lysozyme structure. The concentrated PILs with longer alkyl chain lengths, i.e., over 10 mol% butyl-, 5 mol% hexyl- and 1 mol% octylammonium cations, possessed liquid nanostructures. This detrimentally interfered with solvent subtraction, and the more structured PIL solutions prevented quantitative SAXS analysis of lysozyme structure. The radius of gyration (R-g) of lysozyme in the less structured aqueous PIL solutions showed little change with up to 10 mol% of PIL. Kratky plots, SREFLEX models, and FTIR data showed that the protein conformation was maintained at a low PIL concentration of 1 mol% and lower when compared with the buffer solution. However, 50 mol% EAN and 5 mol% HAN significantly increased the R-g of lysozyme, indicating unfolding and aggregation of lysozyme. The hydrophobic interaction and liquid nanostructure resulting from the increased cation alkyl chain length in HAN likely becomes critical. The impact of HAN and OAN, particularly at high concentrations, on lysozyme structure was further revealed by FTIR. This work highlights the negative effect of a long alkyl chain length and high concentration of PILs on lysozyme structural stability. (AU)

• FAPESP's process: 18/50009-8 - Understanding the molecular interactions between ionic liquids and biopharmaceuticals: the key for a proper design of downstream processing
• Grantee: Jorge Pereira
• Support Opportunities: Regular Research Grants
• FAPESP's process: 20/14144-8 - Purification and encapsulation of bacteriocins from probiotic bacteria for applications in animal protein production systems
• Grantee: Nathalia Vieira Porphirio Veríssimo
• Support Opportunities: Scholarships in Brazil - Post-Doctoral