Targeting the solvent accessible surface area (SASA) in proteins

DELFINO J.M; BERNAR, EM; GÓMEZ G.E

Castellón

Congreso; 6th International Iberian Biophysics Congress and X Iberoamerican Congress of Biophysics; 2018

The hydrophobic effect is widely regarded as the major driving force behind folding and interactions in proteins. Consequently, SASA and its counterpart, the excluded volume, become topographic descriptors of a given folding state. However, these fundamental signatures elude straightforward experimental scrutiny. With the aim at evaluating the size, location and nature of SASA, we introduced a photochemical approach mimicking the role played by the water solvent. The rather indiscriminate reaction of diazirine (DZN), a minute CNN heterocycle, with the polypeptide chain leaves a methylated trace as the new observable. Quantitative assessment of the extent of modification is possible through metrics based on radiochemical or mass spectrometric techniques. Unlike the former, the latest multidimensional NMR advance does not demand cleavage into peptides or amino acids to pinpoint the locale of labeling. A combined analysis, including 1H-NMR, 1H-15N-HSQC and 1H-13C-HSQC spectra, was carried out on the paradigm of E. coli thioredoxin (TRX). Methylation of amino acid side-chains represents the prevailing phenomenon, bearing dissimilar influence on backbone amide environments. Furthermore, inner surfaces such as those delimiting the boundaries of cavities, become prime targets for DZN. The fully consistent picture emerging from this footprint on solvent exposure illuminates relevant traits of protein conformation.