High-resolution crystal structures of villin headpiece and mutants with reduced F-actin binding activity.

Villin-type headpiece domains are approximately 70 amino acid modular motifs found at the C terminus of a variety of actin cytoskeleton-associated proteins. The headpiece domain of villin, a protein found in the actin bundles of the brush border epithelium, is of interest both as a compact F-actin binding domain and as a model folded protein. We have determined the high-resolution crystal structures of chicken villin headpiece (HP67) at 1.

The dual-function disabled-1 PTB domain exhibits site independence in binding phosphoinositide and peptide ligands.

While typical intracellular protein modules have only one ligand-binding site, there are rare examples of single modules that bind two different ligands at distinct binding sites. Here we present a detailed mutational and energetic analysis of one such domain, the phosphotyrosine binding (PTB) domain of Disabled-1 (Dab1), which binds to both peptide and phosphoinositide (PI) ligands simultaneously at structurally distinct binding sites. Through the techniques of isothermal titration calorimetry (ITC), analysis of Dab1 PTB domain mutants, and nuclear magnetic resonance (NMR), we have evaluated the characteristics of binding of the Dab1 PTB domain to various peptide and PI ligands.

The NMR structure of dematin headpiece reveals a dynamic loop that is conformationally altered upon phosphorylation at a distal site.

Dematin (band 4.9) is found in the junctional complex of the spectrin cytoskeleton that supports the erythrocyte cell membrane. Dematin is a member of the larger class of cytoskeleton-associated proteins that contain a modular "headpiece" domain at their extreme C termini.

Nuclear magnetic resonance structure of a prototype Lin12-Notch repeat module from human Notch1.

Notch1 is a member of a conserved family of large modular heterodimeric type 1 transmembrane receptors that control differentiation in multicellular animals. Receptor maturation is accompanied by a furin-dependent cleavage that converts the Notch1 precursor polypeptide into a heterodimer consisting of an extracellular ligand-binding subunit (NEC) and a transmembrane signaling subunit (NTM). Binding of a physiologic ligand to NEC induces signaling by triggering additional proteolytic cleavages in NTM, which allow its intracellular region to translocate to the nucleus where it participates in a transcriptional activation complex.

The role of aromatic residues in the hydrophobic core of the villin headpiece subdomain.

Small autonomously folding proteins are of interest as model systems to study protein folding, as the same molecule can be used for both experimental and computational approaches. The question remains as to how well these minimized peptide model systems represent larger native proteins. For example, is the core of a minimized protein tolerant to mutation like larger proteins are? Also, do minimized proteins use special strategies for specifying and stabilizing their folded structure? Here we examine these questions in the 35-residue autonomously folding villin headpiece subdomain (VHP subdomain).