Concentration-dependent changes to diffusion and chemical shift of internal standard molecules in aqueous and micellar solutions

Abstract

Sodium 4,4-dimethyl-4-silapentane-1-sulfonate (DSS) is the most widely accepted internal standard for protein NMR studies in aqueous conditions. Since its introduction as a reference standard, however, concerns have been raised surrounding its propensity to interact with biological molecules through electrostatic and hydrophobic interactions. While DSS has been shown to interact with certain proteins, membrane protein studies by solution-state NMR require use of membrane mimetics such as detergent micelles and, to date, no study has explicitly examined the potential for interaction between membrane mimetics and DSS. Consistent with its amphipathic character, we show DSS to self-associate at elevated concentrations using pulsed field gradient-based diffusion NMR measurements. More critically, DSS diffusion is significantly attenuated in the presence of either like-charged sodium dodecyl sulfate or zwitterionic dodecylphosphocholine micelles, the two most commonly used detergent-based membrane mimetic systems used in solution-state NMR. Binding to oppositely charged dodecyltrimethylammonium bromide micelles is also highly favourable. DSS-micelle interactions are accompanied by a systematic, concentration- and binding propensity-dependent change in the chemical shift of the DSS reference signal by up to 60 ppb. The alternative reference compound 4,4-dimethyl-4-silapentane-1-ammonium trifluoroacetate (DSA) exhibits highly similar behaviour, with reversal of the relative magnitude of chemical shift perturbation and proportion bound in comparison to DSS. Both DSS and DSA, thus, interact with micelles, and self-assemble at high concentration. Chemical shift perturbation of and modulation of micellar properties by these molecules has clear implications for their use as reference standards.

NMR Precision Metabolomics: Dynamic Peak Sum Thresholding and Navigators for Highly Standardized and Reproducible Metabolite Profiling of Clinical Urine Samples

“…DSS is the most widely accepted internal standard for NMR studies. However, previous reports have demonstrated it has a propensity to interact with biological molecules through electrostatic and hydrophobic interactions [43]. This has led some to suggest that the compound dimethyl-4-silapentane-1-ammonium trifluoroacetate (DSA), which is less likely to bind to proteins, should be used as an alternative reference [44].…”

“…This has led some to suggest that the compound dimethyl-4-silapentane-1-ammonium trifluoroacetate (DSA), which is less likely to bind to proteins, should be used as an alternative reference [44]. However, DSA is more expensive than DSS, and recently both DSS and DSA were shown to interact with micelles [43]. Here, we exploited the natural tendency of DSS to bind proteins to demonstrate that the peak width of DSS can be used to monitor samples for protein contamination.…”

Diastereomers of the anticancer peptide CIGB-300 with altered b-turn structures

“…For simplification of the aromatic region in the spectra, the respective peptide isomers were dissolved in D2O for 1D 1 H-NMR comparisons. In addition, 0.03% DSS was added to all samples for chemical shift referencing (Morash et al 2018), and the pH was maintained between 3.2 and 3.5. The measurements were conducted using a Bruker US2 800 Avance III spectrometer operating at a proton frequency of 800.23 MHz using a TXO cryoprobe.…”