Expedited Nuclear Magnetic Resonance Assignment of Small- to Medium-Sized Molecules with Improved HSQC−CLIP−COSY Experiments

Abstract

Resonance assignment is a pivotal step for any NMR analysis, such as structure elucidation or investigation of protein-ligand interactions. Both 1 H-13 C HSQC and 1 H-1 H COSY 2D experiments are invaluable for 1 H NMR assignment, by extending the high signal dispersion of 13 C chemical shifts onto the 1 H resonances, and by providing a high amount of through-bond 1 H-1 H connectivity information, respectively. The recently introduced HSQC-CLIP-COSY method combines these two experiments, providing COSY correlations along the high-resolution 13 C dimension with clean inphase multiplets. However, two experiments need to be recorded to unambiguously identify COSY cross-peaks. Here, we propose novel variants of the HSQC-CLIP-COSY pulse sequence that edit cross-peak signs so that direct HSQC responses can be distinguished from COSY relay peaks, and/or the multiplicities of the 13 C nuclei are reflected, allowing assignment of all peaks in just a single experiment. The advanced HSQC-CLIP-COSY variants have the potential to accelerate and simplify the NMR structure elucidation process of both synthetic and natural products, and to become valuable tools for high-throughput computer-assisted structure determination.

2-Acetamido-2-deoxy-d-glucono-1,5-lactone Sulfonylhydrazones: Synthesis and Evaluation as Inhibitors of Human OGA and HexB Enzymes

“…Chemical shifts are referenced to TMS as an internal reference ( 1 H) or to residual solvent signals ( 13 C). 2D 1 H- 13 C HSQC-CLIP-COSY experiment with HSQC vs. COSY peak sign editing [26] and 2D 1 H-1 H EASY ROESY experiment [19] with 300 ms mixing time were carried out for the unambiguous structure elucidation of compound 5d on a Bruker Avance Neo 700 MHz NMR spectrometer (Bruker, Karlsruhe, Germany) equipped with a Prodigy TCI cryoprobe. Mass spectra were recorded with MicroTOF-Q type Qq-TOF MS and maXis II UHR ESI-QTOF MS (Bruker Daltonik, Bremen, Germany) instruments in the positive ion mode with the electrospray ionization or atmospheric pressure chemical ionization technique.…”

Primary Structure of Glycans by NMR Spectroscopy

“…The CLIP module has been utilized in CLIP-COSY relayed, 1 H, 13 C-HSQC-CLIP-COSY and 1 H, 13 C-HSQC-CLIP-COSY relayed experiments applied to tri- and pentasaccharides for NMR resonance assignments . Furthermore, the possibility to obtain 1 H, 13 C-HSQC-CLIP-COSY spectra, where cross-peaks have different signs depending on whether they originate from direct HSQC responses or from COSY cross-peaks, have been described as an additional improvement …”

Synthesis of a Heparinoid Pentasaccharide Containing l-Guluronic Acid Instead of l-Iduronic Acid with Preserved Anticoagulant Activity

“…The complete and unambiguous 1 H and 13 C NMR resonance assignment of pentasaccharide 8 was achieved with the aid of classical ( 1 H− 1 H COSY, TOCSY, 1 H− 13 C HSQC, and HMBC) and advanced ( 1 H− 1 H EASY ROESY, 36 1 H− 13 C HSQC-CLIP-COSY, 37 and NORD HSQC-TOCSY 38 ) 2D NMR experiments. As the ring conformation of unit G has been suggested as a major determinant of biological activities, we planned to study the conformation of the L-guluronic acid unit of 8.…”