Identification of the PXW Sequence as a Structural Gatekeeper of the Headpiece C-terminal Subdomain Fold
| Autor: | Daisy Dewitte, David Bourry, Frans Borremans, José C. Martins, Stefaan Rossenu, Christophe Ampe, Marleen Van Troys, Joël Vandekerckhove, W Vermeulen, Marc Goethals |
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| Rok vydání: | 2006 |
| Předmět: |
Models
Molecular Protein Folding Proline Stereochemistry Protein subunit Molecular Sequence Data Stacking macromolecular substances Evolution Molecular Protein stability Neurofilament Proteins Structural Biology Animals Humans Amino Acid Sequence Actin-binding protein Nuclear Magnetic Resonance Biomolecular Molecular Biology Conserved Sequence biology Chemistry Lysine Microfilament Proteins Titrimetry Tryptophan Blood Proteins Hydrogen-Ion Concentration Phosphoproteins Actins Peptide Fragments Protein Structure Tertiary Crystallography Amino Acid Substitution Mutation biology.protein Sequence motif Villin Chickens Protein Binding |
| Zdroj: | Journal of Molecular Biology. 359:1277-1292 |
| ISSN: | 0022-2836 |
| DOI: | 10.1016/j.jmb.2006.04.042 |
| Popis: | The HeadPiece (HP) domain, present in several F-actin-binding multi-domain proteins, features a well-conserved, solvent-exposed PXWK motif in its C-terminal subdomain. The latter is an autonomously folding subunit comprised of three alpha-helices organised around a hydrophobic core, with the sequence motif preceding the last helix. We report the contributions of each conserved residue in the PXWK motif to human villin HP function and structure, as well as the structural implications of the naturally occurring Pro to Ala mutation in dematin HP. NMR shift perturbation mapping reveals that substitution of each residue by Ala induces only minor, local perturbations in the full villin HP structure. CD spectroscopic thermal analysis, however, shows that the Pro and Trp residues in the PXWK motif afford stabilising interactions. This indicates that, in addition to the residues in the hydrophobic core, the Trp-Pro stacking within the motif contributes to HP stability. This is reinforced by our data on isolated C-terminal HP subdomains where the Pro is also essential for structure formation, since the villin, but not the dematin, C-terminal subdomain is structured. Proper folding can be induced in the dematin C-terminal subdomain by exchanging the Ala for Pro. Conversely, the reverse substitution in the villin C-terminal subdomain leads to loss of structure. Thus, we demonstrate a crucial role for this proline residue in structural stability and folding potential of HP (sub)domains consistent with Pro-Trp stacking as a more general determinant of protein stability. |
| Databáze: | OpenAIRE |
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