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Publication Abstract from PubMed

Incorporating metal cofactors into computationally designed protein scaffolds provides a versatile route to novel protein functions, including the potential for new-to-nature enzyme catalysis. However, a major challenge in protein design is to understand how the scaffold architecture influences conformational dynamics. Here, we characterized structure and dynamics of a modular de novo scaffold with flexible inter-domain linkers. Three rationally engineered variants with different metal specificity were studied by combining X-ray crystallography, NMR spectroscopy, and molecular dynamics simulations. The lanthanide-binding variant was initially trapped in an inactive conformational state, which impaired efficient metal coordination and cerium-dependent photocatalytic activity. Stabilization of the active conformation by AI-guided sequence optimization using ProteinMPNN led to accelerated lanthanide binding and a 10-fold increase in k(cat)/K(m) for a photoenzymatic model reaction. Our results suggest that modular scaffold architectures provide an attractive starting point for de novo metalloenzyme engineering and that ProteinMPNN-based sequence redesign can stabilize desired conformational states.

Modular protein scaffold architecture and AI-guided sequence optimization facilitate de novo metalloenzyme engineering.,Wagner Egea P, Delhommel F, Mustafa G, Leiss-Maier F, Klimper L, Badmann T, Heider A, Wille I, Groll M, Sattler M, Zeymer C Structure. 2025 Nov 5:S0969-2126(25)00397-1. doi: 10.1016/j.str.2025.10.010. PMID:41197620^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.