FIGURE 4.

S3 and DBL3x binding to CSA oligosaccharide observed by STD NMR. A–C, STD spectrum (C) of 60 μm S3 protein without CSA oligosaccharide at 20 °C, 1024 scans, obtained by subtraction (C = B − A) of the on- (A) from the off- (B) saturation spectra. Present in solution are some low molecular weight contaminants (asterisk), but note that their signals are completely cancelled in the difference spectrum in C. D, 1D ¹H NMR spectrum of CSA oligosaccharide (20 °C, 16 scans, no water suppression). Residual signals from chondroitin-6-O-sulfate are labeled with x. Due to overlap, only selected resonances are labeled. E, STD spectrum of 1:100 molar ratio of S3 and CSA oligosaccharide. F, STD spectrum of 1:100 molar ratio of DBL3x and CSA oligosaccharide at 20 °C using 1024 scans each. G, same as E but the STD data were collected without water suppression at 10 °C to observe the STD signals for the GalNAc-H4, GalNAc-H1, and GlcA-H1 resonances that were obscured by water suppression in E. HDO, residual partially deuterated water.