Elucidation of Gd(III)-Ligand and Gd(III)-Protein Complexes for Development of Gd(III) Contrast Agents Using High-Frequency EPR Spectroscopy

/Abstract Gd3+-based contrast agent (GBCA) enables enhanced MRI images for visualizing internal organs, tissues, and blood vessels. While GBCA is primarily considered safe, trace amounts of Gd have been detected in the bones and brains of patients after multiple administration of GBCA. Additionally, a life-threatening but rare nephrogenic systemic fibrosis disease can occur in patients with chronic kidney disease after exposure to GBCA. Currently, the use of GBCA is restricted against patients who are at risk. Hence, developing a new GBCA is of high interest from the perspective of contrasting capability and safety. However, such development requires the ability to diagnose GBCA stability both in vitro and in vivo. In this proposal, we will develop a unique high-powered 263 GHz Electron Paramagnetic Resonance (EPR) spectrometer that will enable sensitive detection and highly- resolved characterization of the Gd3+ coordination environment in both test tubes and cells. The new spectrometer will be at the forefront of Gd3+-based magnetic resonance, paving the way for describing the physical chemistry of Gd3+. We will also establish the EPR spectral features associated with Gd3+ bound to its chelating carrier or dissociated Gd3+ that has interacted with endogenous proteins. Specifically, we will use a class of lanthanide-binding peptides known as Lanthanide-Binding Tag (LBT3) and a recently discovered picomolar lanthanide-binding protein, LanM, which has invigorated the field of lanthanide biochemistry. The high- frequency Gd3+ EPR spectra contain zero-field splitting features that serve as the spectral 'fingerprint' of different Gd3+-protein interactions, crucial for diagnosing either ligand-bound or protein-bound Gd3+. Finally, we will use LanM and engineered LBT3 as candidates for new GBCAs. The picomolar affinities of these proteins, which have been exploited for commercial lanthanide extraction strategies, are also highly promising for carriers of GBCA. This research plan aims to integrate EPR methodologies to gain insight into the structural features of Gd3+-ligand and Gd3+-protein interactions for future GBCA development. The long-term goal is to develop a method that can identify the different Gd3+ complexes, either to assess the stability of GBCA in a biological context or to create new GBCAs with new coordination environments. The fellowship will support the applicant in gaining expertise in microwave engineering and protein engineering, preparing him for a well-rounded career as an investigator at the front line of magnetic resonance spectroscopy and lanthanide-related structural biology. Project Number: 1F32GM159401-01A1 | Fiscal Year: 2026 | NIH Institute/Center: National Institute of General Medical Sciences (NIGMS) | Principal Investigator: Zikri Hasanbasri | Institution: UNIVERSITY OF CALIFORNIA AT DAVIS, DAVIS, CA | Award Amount: $79,348 | Activity Code: F32 | Study Section: Special Emphasis Panel[ZRG1 F05-D (21)] View on NIH RePORTER: https://reporter.nih.gov/project-details/11314892