# Development of LED-Assisted NMR Technologies for the Atomic-Resolution Analysis of Medically Relevant Biomolecules in Solution at Submicromolar Concentration

> **NIH NIH R01** · UNIVERSITY OF WISCONSIN-MADISON · 2024 · $488,401

## Abstract

Project Summary
The goal of this R01 renewal application is to further develop the LED-enhanced NMR technology (LC-photo-
CIDNP) that was established during the prior cycle of funding, and extend the method to the facile and ultra-
rapid 1D-to-3D NMR spectroscopy of proteins at sub-micromolar concentration. We will focus on NMR studies
in solution and will target folded, unfolded and intrinsically disordered proteins in either buffered solution or cell-
like media. We will accomplish the above goals within three steps. First (Specific Aim #1), we will incorporate a
tryptophan (Trp) isotopolog bearing a quasi-isolated 1H-13C spin pair (QISP) within soluble proteins to
achieve unprecedented NMR sensitivity for the detection of solvent-exposed Trp in proteins at nanomolar and
sub-nanomolar levels. We will then employ the above technology in combination with field-cycling to achieve
further NMR sensitivity enhancements. Second (Specific Aim #2) we will extend LC-photo-CIDNP to amino
acids other than Trp and Tyr within proteins. This goal will be accomplished via through-space and through-
bond polarization transfer methodologies. Third (Specific Aim #3), we will extend LC-photo-CIDNP to higher-
dimensionality (>2D) NMR spectroscopy by developing novel 3D (and possibly 4D) 1H,13C heteronuclear
spectroscopy pulse sequences tailored to the analysis of side-chain and backbone 1H-13C resonance pairs.
This effort will include non-uniform-sampling (NUS) data collection schemes. We will then combine theoretical
calculations and experiments to develop better LC-photo-CIDNP dyes with optimized g-factor values and long
photoexcited-state lifetimes, for optimal LC-photo-CIDNP data collection. We will also exploit the peculiar field
dependence of LED-enhanced NMR and implement 2D LC-photo-CIDNP on benchtop NMR spectrometers.
Finally, we will test the success of the improved LC-photo-CIDNP technologies developed in this work by
studying the interaction of an aggregation-prone client protein (SH3 variant) with the Hsp70 molecular
chaperone at sub-micromolar concentration.

## Key facts

- **NIH application ID:** 10902100
- **Project number:** 5R01GM125995-06
- **Recipient organization:** UNIVERSITY OF WISCONSIN-MADISON
- **Principal Investigator:** Silvia Cavagnero
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $488,401
- **Award type:** 5
- **Project period:** 2018-09-08 → 2027-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10902100

## Citation

> US National Institutes of Health, RePORTER application 10902100, Development of LED-Assisted NMR Technologies for the Atomic-Resolution Analysis of Medically Relevant Biomolecules in Solution at Submicromolar Concentration (5R01GM125995-06). Retrieved via AI Analytics 2026-07-28 from https://api.ai-analytics.org/grant/nih/10902100. Licensed CC0.

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