A series of NMR supersequences are presented for the time‐efficient structure characterisation of small molecules in the solution state. These triplet sequences provide HMBC, HSQC, and one homonuclear correlation experiment of choice according to the NMR by Ordered Acquisition using 1H detection principle. The experiments are demonstrated to be compatible with non‐uniform sampling schemes and may be acquired and processed under full automation.

Galectin-3 is crucial to many physiological and pathological processes. The generally accepted dogma is that galectins function extracellularly by binding specifically to β(1➔4)-galactoside epitopes on cell surface glycoconjugates. Here, we used crystallography and NMR spectroscopy to demonstrate that negatively charged homogalacturonans (HG, linear polysaccharides of α(1➔4)-linked-D-galacturonate (GalA)) bind to the galectin-3 carbohydrate recognition domain. The HG carboxylates at the C6 positions in GalA rings mandate that this saccharide bind galectin-3 in an unconventional, “topsy-turvy” orientation that is flipped by about 180o relative to that of the canonical β-galactoside lactose. In this binding mode, the reducing end GalA β-anomer of HGs takes the position of the non-reducing end galactose residue in lactose. This novel orientation maintains interactions with the conserved tryptophan and seven of the most crucial lactose-binding residues, albeit with different H-bonding interactions. Nevertheless, the HG molecular orientation and new interactions have essentially the same thermodynamic binding parameters as lactose. Overall, our study provides structural details for a new type of galectin-sugar interaction that broadens glycospace for ligand binding to Gal-3 and suggests how the lectin may recognize other negatively-charged polysaccharides like glycoaminoglycans (e.g. heparan sulfate) on the cell surface. This discovery impacts on our understanding of galectin-mediated biological function.

Abstract13C nuclear magnetic resonance (NMR) is traditionally considered an insensitive technique, requiring long acquisition times to measure dilute functionalities on large polymers. With the introduction of cryoprobes and better electronics, sensitivity has improved in a way that allows measurements to take less than 1/20th the time that they previously did. Unfortunately, a high Q‐factor with cryoprobes creates baseline curvature related to acoustic ringing that affects quantitative NMR analyses. Manual baseline correction is commonly used to compensate for the baseline roll, but it is a time‐intensive process. The outcome of manual baseline correction can vary depending on processing parameters, especially for complicated spectra. Additionally, it can be challenging to distinguish between broad peaks and baseline rolls. A new anti‐ring pulse sequence (zgig_pisp) was previously reported to improve on the incumbent single pulse experiment (zgig). The original report presented limited comparison data with 13C NMR, but a thorough validation is needed before broader implementation can be considered. In this work, we report the round‐robin testing and comparison of zgig_pisp and zgig pulse sequences. During the testing phase, we found that zgig_pisp is practically equivalent to zgig to ±2% for the majority of integrals examined. Additionally, a short broadband inversion pulse (BIP) was demonstrated as an alternative to the originally reported adiabatic CHIRP shaped pulse. The zgig_pisp pulse sequence code for Bruker spectrometers is also simplified.

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