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Overlay of 1H-15N SOFAST spectra of the middle Ub in all-natural mixed K33,K11-linked Ub3 (red) and in blue, (a) WT Ub, (b) the proximal Ub of K33-linked Ub2. The spectral differences between residues in the middle Ub unit of K33,K11-linked Ub3 and (c) WT monoUb and (d) proximal Ub of K33-linked Ub2 are quantified as amide chemical shift perturbations (CSPs). Residues with significant CSPs are indicated on the spectra in panels a and b.
