Kate Hadley (@ka-hadley.bsky.social)

Metallo-coiled Coil Stabilization via Chemical Cross-Linking: Implications for Gd(III)-Based MRI Contrast Agents Coiled coils are a versatile class of protein-inspired metal-binding ligands with well-defined sequence-to-structure relationships and high synthetic accessibility, making them promising tunable ligands for applications spanning biomedical imaging, catalysis, and material science. However, their practical utility is often limited by poor stability, particularly under physiological conditions. Herein, we introduce a covalent interhelical isopeptide cross-linking strategy that significantly enhances the thermal, thermodynamic, kinetic, and proteolytic stability of lanthanide-bound coiled coils designed for use as MRI contrast agents. Biophysical analysis, including CD, native MS, HPLC, and fluorescence assays, reveal that introduction of a single cross-linking layer promotes structural organization, driving the peptide from an unfolded to a well-folded state. Cross-linking leads to a remarkable 2.5 × 108-fold enhancement in metal-binding affinity, along with greater kinetic stability and resistance to proteolytic degradation compared to the non-cross-linked analogue. 1H relaxometric studies and molecular dynamics simulations reveal that this class of potential MRI contrast agents operates via a second-sphere water coordination mechanism. Optimized cross-linking improves MRI efficacy by ∼30% at clinically relevant field strengths, highlighting its potential as a design principle for next-generation MRI contrast agents. Beyond MRI, these findings underscore the broader potential of covalent cross-linking strategies for enhancing the stability and functionality of metallo-coiled coils, expanding their utility across diverse applications. https://pubs.acs.org/doi/10.1021/jacs.5c13620