Dissecting neurofilament tail sequence-phosphorylation-structure relationships with multicomponent reconstituted...

Area of Science:

• Biophysics
• Cell Biology
• Neuroscience

Background:

• Neurofilaments (NFs) are key axonal cytoskeleton components with intrinsically disordered tail domains.
• The conformation and function of NF tail brushes, influenced by charge and phosphorylation, are not fully understood.

Purpose of the Study:

• To investigate how neurofilament tail protein constructs (NFL, NFM, NFH) form protein brushes.
• To determine the impact of composition, phosphorylation, and charge patterns on brush conformation and morphology.

Main Methods:

• Grafting recombinant NF tail proteins (NFL, NFM, NFH) onto surfaces to create defined protein brushes.
• Atomic force microscopy (AFM) to measure brush height and morphology.
• Self-consistent field theory (SCFT) modeling to predict and analyze brush structures.

Main Results:

• Brush height depends monotonically but not always linearly on composition.
• NFM brushes are highly extended; NFH brushes are compact despite phosphorylation.
• SCFT predicts multilayer morphologies for NFM and phosphorylated NFH brushes, driven by N-terminal charges in NFH and charge segregation in NFM.

Conclusions:

• NF-M plays a role in the dynamic range of NF brush conformation.
• NFH tail N-terminal charges and C-terminal regions influence multilayer formation and height.
• Disordered protein sequence contributions to interfacial conformation can be dissected using this platform.