The Effect of Dimethyl Sulfoxide on the Lysozyme Unfolding Kinetics, Thermodynamics, and Mechanism

Abstract

The thermal stability of proteins in the presence of organic solvents and the search for ways to increase this stability are important topics in industrial biocatalysis and protein engineering. The denaturation of hen egg-white lysozyme in mixtures of water with dimethyl sulfoxide (DMSO) with a broad range of compositions was studied using a combination of differential scanning calorimetry (DSC), circular dichroism (CD), and spectrofluorimetry techniques. In this study, for the first time, the kinetics of unfolding of lysozyme in DMSO–water mixtures was characterized. In the presence of DMSO, a sharp decrease in near-UV CD and an increase in the fluorescence signal were observed at lower temperatures than the DSC denaturation peak. It was found that differences in the temperatures of the CD and DSC signal changes increase as the content of DMSO increases. Changes in CD and fluorescence are triggered by a break of the tertiary contacts, leading to an intermediate state, while the DSC peak corresponds to a subsequent complete loss of the native structure. In this way, the commonly used two-state model was proven to be unsuitable to describe the unfolding of lysozyme in the presence of DMSO. In kinetic studies, it was found that even high concentrations of DMSO do not drastically change the activation energy of the initial stage of unfolding associated with a disruption of the tertiary structure, while the enthalpy of denaturation shows a significant dependence on DMSO content. This observation suggests that the structure of the transition state upon unfolding remains similar to the structure of the native state.

Mix and Shake: A Mild Way to Drug-Loaded Lysozyme Nanoparticles

“…This highlights that lysozyme pretreated with DMSO supports the formation of better-defined nanoparticles (Figures , S2, and S3). While there is always a concern that lysozyme could denature in DMSO , (although in our case it does not), there is an advantage of using small amounts of organic solvent as it helps to dissolve protein and drug and they can both interact with fructose. , To explore if the ability of lysozyme to form well-defined nanoparticles is unique, albumin and fibrinogen were, in addition, tested using similar procedures as that for Lys-Fru-CCM particles (for details, see Materials and Methods, Protocol 2). Only protocol 2 was used here, as albumin is not DMSO soluble and fibrinogen tends to denature.…”

“…and plants, serving as part of the innate immune response by protecting against bacteria and viruses. , Lysozyme can disrupt the bacterial wall due to its positive charge, and, in combination with antibiotics, can partially decrease the drug resistance due to a synergistic effect. , Furthermore, lysozyme is a monomeric protein, and its three-dimensional structure is stabilized by only four disulfide linkages. As a result, the denaturation process of lysozyme can be reversed relatively easily, − endowing lysozyme with relatively high stability against denaturation. Lysozyme has already been used in the delivery of hydrophobic drugs. − Due to the smaller size, lysozyme with conjugated drugs accumulates less likely in the kidney .…”

Interaction investigation and phase transition of carrageenan/lysozyme complex system

“…As the temperature of the system increased, the fluorescence spectrum peaks significantly decreased, which led to corresponding changes in the composite system. This was because heating caused greater changes in the secondary structure of Ly, and the hydrophobic interaction inhibited the formation of Ly/CRG complexes to a weaker extent (Magsumov et al, 2019). As the temperature increased, the content of α-helix in Ly decreased, and the content of random coils increased.…”

Improving the reproducibility of size distribution of protein crystals produced in continuous slug flow crystallizer operated at short residence time

“…Organic solvents are good for large scale precipitation since they can be recycled efficiently, yet it is well-known that the organic solvents generally denature protein/peptides [115,151,195,196]. When used in high concentration or high temperature, the Dimethyl Sulfoxide (DMSO) might lead to the denaturation of lysozyme via disrupting the tertiary structure of the molecules that leads to unfolding [197]. Besides, different organic solvents were found to affect the particle size, zeta potential, secondary structure, thermodynamic properties (Tm values), and the biological activity of lysozyme differently [198].…”