Detection of 15N-labeled Metabolites in Microbial Extracts using AI-Designed Broadband pulses for 1H, 15N Heteronuclear NMR Spectroscopy

Abstract

Approximately 40% of bacterial and mammalian metabolites contain nitrogen-based chemical moieties such as amides, amines, and imines. The identification and quantification of these groups via 2D 1H-15N heteronuclear NMR spectroscopy have broadened the catalog of NMR-detected metabolites. However, these NMR experiments necessitate broadband ra-diofrequency (RF) pulses for inversion and refocusing operations to encompass the full range of 15N chemical shifts, a chal-lenge that becomes increasingly apparent at high and ultra-high magnetic fields. Here, we show that a newly AI-designed broadband 15N universal 180o pulse for both inversion and refocusing incorporated in the 2D 1H, 15N heteronuclear single quantum coherence (2D 1H-15N BB-HSQC) experiment significantly enhances spectral sensitivity. We demonstrate the ad-vantage of the new technique by analyzing the crude extract of Micromonospora sp. WMMC264, a microbial strain that produces siderophores for iron absorption from the environment. The implementation of the AI-designed pulse in the 2D 1H-15N BB-HSQC experiment will contribute to advancing the analysis of nitrogen-containing metabolites in biological fluids and cell extracts.