Practical considerations for the measurement of near-surface electrostatics based on solvent paramagnetic relaxation enhancements.

Electrostatic interactions can play important roles in regulating various biological processes. Quantifying surface electrostatics of biomolecules is, therefore, of significant interest. Recent advances in solution NMR spectroscopy have enabled site-specific measurements of de novo near-surface electrostatic potentials (ϕENS) based on a comparison of solvent paramagnetic relaxation enhancements generated from differently charged paramagnetic co-solutes with similar structures. Although the NMR-derived near-surface electrostatic potentials have been shown to agree with theoretical calculations in the context of folded proteins and nucleic acids, such benchmark comparisons may not always be possible, particularly in cases where high-resolution structural models are lacking, such as in the study of intrinsically disordered proteins. Cross-validation of ϕENS potentials can be achieved by comparing values obtained using three pairs of paramagnetic co-solutes, each with a different net charge. Notably we have found cases where agreement of ϕENS potentials between the three pairs is poor and herein we investigate the source of this discrepancy in some detail. We show that for the systems considered here ϕENS potentials obtained from cationic and anionic co-solutes are accurate and that the use of paramagnetic co-solutes with different structures can be a viable option for validation, although the optimal choice of paramagnetic compounds depends on the system of interest.