Ultrafast Single‐Scan 2D NMR Spectroscopic Detection of a PHIP‐Hyperpolarized Protease Inhibitor

Abstract

Two‐dimensional NMR spectroscopy is one of the most important spectroscopic tools for the investigation of biological macromolecules. However, due to the low sensitivity of NMR spectroscopy, it takes usually from several minutes to many hours to record such spectra. Here, the possibility of detecting a bioactive derivative of the sunflower trypsin inhibitor‐1 (SFTI‐1), a tetradecapeptide, by combining parahydrogen‐induced polarization (PHIP) and ultrafast 2D NMR spectroscopy is shown. The PHIP activity of the inhibitor was achieved by labeling with O‐propargyl‐l‐tyrosine. In 1D PHIP experiments a signal enhancement of a factor of approximately 1200 compared to standard NMR was found. This enhancement permits measurement of 2D NMR correlation spectra of low‐concentrated SFTI‐1 in less than 10 seconds, employing ultrafast single‐scan 2D NMR detection. As experimental examples PHIP‐assisted ultrafast single‐scan TOCSY spectra of SFTI‐1 are shown.

Parahydrogen‐Induced Polarization of Amino Acids

“…Signals marked with an asterisk (*) are impurities (adapted from Sauer et al [68] . and Kiryutin et al [67] …”

Sunflower Trypsin Inhibitor‐1 (SFTI‐1): Sowing Seeds in the Fields of Chemistry and Biology

“…The labelled molecule was immediately analysed by NMR spectroscopy using a PHIP irradiation sequence, which resulted in a 70‐fold enhancement of the 1 H NMR signal [105] . Further optimisation of the hyperpolarisation conditions resulted in a 1200‐fold enhancement of the 1 H signal, thereby enabling acquisition of 2D 1 H NMR spectra for an SFTI‐1 analogue in less than 10 seconds [106] . This technique has considerable potential and represents a viable method for the in situ analysis of biomolecules by NMR spectroscopy.…”

Parawasserstoff‐induzierte Polarisation von Aminosäuren

“…[75] Die Reproduzierbarkeit dieses Aufbaus ermçglichte PHIP-verstärkte 2D-NMR-Experimente,d ie aufgrund von Schwankungen in der Polarisationsausbeute bei manueller Durchführung normalerweise schwierig sind. Um dieses Problem anzugehen, wurden ultraschnelle 2D-NMR-Methoden vorgeschlagen (Abbildung 6), [67,76,77] die von der Gruppe von Frydman entwickelt wurden. [78][79][80][81] Ultraschnelle 2D-NMR kombiniert Ideen aus der NMR und MRT, z.…”