A family portrait of lanmodulin selectivity for enhanced rare-earth separations

Proteins offer a molecular design space to create bespoke ligands for the separation of critical metals, like rare earth elements (REs). However, data-intensive approaches to fine-tune metalloprotein selectivity are bottlenecked by the low-throughput nature of existing characterization methods. Here we invented an assay called SpyTag-Catcher Immobilization of Lanmodulin for Assaying Metal-Binding Selectivity (SpyCI-LAMBS) to measure metalloprotein selectivity en masse. This 96-format workflow was used to study the selectivity of 621 lanmodulin (LanM) orthologs for 15 REs, revealing eight distinct selectivity profiles based on sequence-to-function analyses. We discovered >200 LanMs with dampened selectivity for low-value LaIII relative to the prototypical LanM. This includes a LanM that can perform a challenging one-stage separation of PrIII from LaIII with up to >99.9 mol% purity and 83% yield. SpyCI-LAMBS is a powerful tool that can rapidly collect high-fidelity selectivity data to inform metal ion separations and machine learning-assisted metalloprotein design.