Article
Received: 2 December 2008; Revised: 19 January 2009; Accepted: 28 January 2009
10.1002/pro.75 About DOI
Redesign of a protein-peptide interaction: Characterization and applications |
| Meredith E. Jackrel 1, Roberto Valverde 2, Lynne Regan 1 2 * |
| 1Department of Chemistry, Yale University, New Haven, Connecticut 06520 2Department of Molecular Biophysics & Biochemistry, Yale University, New Haven, Connecticut 06520 |
| email: Lynne Regan (lynne.regan@yale.edu) |
*Correspondence to Lynne Regan, Yale University, 322 Bass Center, 266 Whitney Ave., New Haven, CT 06511
Funded by:
NIH; Grant Number: GM080515 HFSP; Grant Number: RGP44/2007
| Keywords |
| tetratricopeptide repeat protein (TPR) peptide binding rational design molecular recognition antibody replacements solvent accessible surface area western blotting hydrophobic interactions |
| Abstract |
| The design of protein-peptide interactions has a wide array of practical applications and also reveals insight into the basis for molecular recognition. Here, we present the redesign of a tetratricopeptide repeat (TPR) protein scaffold, along with its corresponding peptide ligand. We show that the binding properties of these protein-peptide pairs can be understood, quantitatively, using straightforward chemical considerations. The recognition pairs we have developed are also practically useful for the specific identification of tagged proteins. We demonstrate the facile replacement of these proteins, which we have termed T-Mods (TPR-based recognition module), for antibodies in both detection and purification applications. The new protein-peptide pair has a dissociation constant that is weaker than typical antibody-antigen interactions, yet the recognition pair is highly specific and we have shown that this affinity is sufficient for both Western blotting and affinity purification. Moreover, we demonstrate that this more moderate affinity is actually advantageous for purification applications, because extremely harsh conditions are not required to dissociate the T-Mod-peptide interaction. The results we present are important, not only because they represent a successful application of protein design but also because they help define the properties that should be sought in other scaffolds that are being developed as antibody replacements. |
Received: 2 December 2008; Revised: 19 January 2009; Accepted: 28 January 2009
| Digital Object Identifier (DOI) |
10.1002/pro.75 About DOI
