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 poten-tials (OENS) based on a comparison of solvent paramagnetic relaxation enhancements generated from dif-ferently 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, particu-larly in cases where high-resolution structural models are lacking, such as in the study of intrinsically dis-ordered proteins. Cross-validation of OENS 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 OENS 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 OENS potentials obtained from cationic and anionic co-solutes are accurate and that the use of paramagnetic co-solutes with dif-ferent structures can be a viable option for validation, although the optimal choice of paramagnetic com-pounds depends on the system of interest.(c) 2023 Elsevier Inc. All rights reserved.