Synergistic Interactions between Mixtures of Ionic and Amphoteric Surfactants Can Enhance Protein Stability

Proteins and surfactants are used in a variety of applications, including the formulation of consumer products such as foods, cosmetics and cleaning agents, and stabilization of biopharmaceuticals. They also find applications in research for tasks such as membrane protein extraction and solubilization and the separation of proteins. The interactions between proteins and surfactants are relevant as they can form complexes that influence protein structure, catalytic activity, aggregation, and binding to surfaces. While the interactions of individual ionic and nonionic surfactants with proteins have been well characterized, detailed studies of protein structure in surfactant mixtures is still an emerging field. We hypothesize that mixed surfactant systems interact differently with proteins based on the surfactant mixture composition, offering the ability to specifically tune interactions between surfactants, micelles, and proteins, impacting protein structure and function. Understanding these interactions is important since they influence product formulation and performance.

In this work, we investigated the interaction of binary mixtures of the anionic surfactant sodium dodecyl sulfate (SDS) and the amphoteric surfactant N,N-dimethyldodecylamine N-Oxide (DDAO) with the model protein β-Lactoglobulin (βLG) using a variety of biophysical techniques including pyrene fluorescence, circular dichroism (CD) and small-angle X-ray scattering (SAXS). The critical micelle concentration (CMC) was unchanged for mixed surfactant samples in the presence of βLG, except above 0.85 mole SDS per mole total surfactant where surfactant aggregates were observed to form below the CMC. This is likely due to surfactant binding to βLG as is observed with neat SDS. Unfolding of βLG was observed in 2 mM DDAO and 1.1 mM SDS of the neat surfactants; however, higher concentrations of the mixed surfactants were required to unfold βLG. Simple linear combination of SAXS scattering curves and predictions of protein secondary structure from CD spectra revealed that βLG structure was most stable in surfactant mixtures rich in DDAO.

We are currently investigating the reason for the improved stability of βLG with the surfactant mixtures. Potential explanations for the improved stability includes decreased surfactant monomer concentrations and the reduced charge of the mixed micelles making interaction with βLG less favorable. Protein refolding from surfactant-unfolded states and the effect of surfactant interactions on enzymatic activity will also be discussed.