A Structural Basis for ¹²⁹ Xe Hyper-CEST Signal in TEM-1 β-Lactamase

Area of Science:

• Biophysics
• Biomolecular Imaging
• Magnetic Resonance Imaging

Background:

• Genetically encoded (GE) contrast agents offer non-invasive MRI visualization of biological processes.
• Hyperpolarized 129Xe and chemical exchange saturation transfer (hyper-CEST) enable ultrasensitive protein detection.
• Existing GE MRI agents include gas vesicles, TEM-1 β-lactamase (bla), and maltose binding protein (MBP).

Purpose of the Study:

• To elucidate the structural basis of hyper-CEST contrast generation by the TEM-1 β-lactamase (bla) protein.
• To characterize the interaction between hyperpolarized 129Xe and the bla protein.

Main Methods:

• Molecular dynamics (MD) simulations to predict Xe binding sites.
• X-ray crystallography to validate predicted Xe binding site location.
• Site-directed mutagenesis to confirm the role of the binding site in CEST contrast.

Main Results:

• MD simulations identified a high-occupancy Xe binding site within the bla protein.
• X-ray crystallography confirmed the Xe binding site location.
• Mutagenesis experiments demonstrated this site is essential for observed CEST contrast.

Conclusions:

• The Xe binding site in bla is structurally validated and crucial for hyper-CEST contrast.
• Understanding protein-Xe interactions advances GE MRI contrast agent development.
• This work provides insights into structure-contrast relationships for biomolecular imaging.