Structural basis for high-affinity actin binding revealed by a β-III-spectrin SCA5 missense mutation

• DOI: 10.1038/s41467-017-01367-w
• Publication Date: 2017-11-01T20:41:03Z
• Authors: Adam W. Avery; Michael E. Fealey; Fengbin Wang; Albina Orlova; Andrew R. Thompson; David D. Thomas; Thomas S. Hays; Edward H. Egelman
• Keywords: Models, Molecular; Binding Sites; Protein Domains; Protein Conformation; Science; Q; Cryoelectron Microscopy; Electron Spin Resonance Spectroscopy; Mutation, Missense; Humans; Spectrin; Article; Actins

AbstractSpinocerebellar ataxia type 5 (SCA5) is a neurodegenerative disease caused by mutations in the cytoskeletal protein β-III-spectrin. Previously, a SCA5 mutation resulting in a leucine-to-proline substitution (L253P) in the actin-binding domain (ABD) was shown to cause a 1000-fold increase in actin-binding affinity. However, the structural basis for this increase is unknown. Here, we report a 6.9 Å cryo-EM structure of F-actin complexed with the L253P ABD. This structure, along with co-sedimentation and pulsed-EPR measurements, demonstrates that high-affinity binding caused by the CH2-localized mutation is due to opening of the two CH domains. This enables CH1 to bind actin aided by an unstructured N-terminal region that becomes α-helical upon binding. This helix is required for association with actin as truncation eliminates binding. Collectively, these results shed light on the mechanism by which β-III-spectrin, and likely similar actin-binding proteins, interact with actin, and how this mechanism can be perturbed to cause disease.