Buttressing ligand-binding pockets in de novo designed NTF2-like domains.

ABSTRACT: NTF2-like proteins are compact α + β fold domains with cone-shaped architectures and internal pockets, making them attractive scaffolds for the de novo design of small-molecule binders and enzymes. However, creating ligand-binding pockets often compromises folding stability, posing a key challenge in de novo protein design. Here, we introduce strategies to stabilize NTF2-like domains while preserving pocket geometry and accessibility. By expanding the hydrophobic core through computationally designed α-helical subdomains or homodimer interfaces buttressing the β-sheet's convex face, we enhance structural stability without blocking pocket access on the concave face. Biochemical, biophysical, and crystallographic analyses confirm that the designed buttressing elements maintain the intended f

SUBMITTER: Nadal M 

PROVIDER: S-EPMC12514758 | biostudies-literature | 2025 Nov

REPOSITORIES: biostudies-literature