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Crystal structure of a consensus-designed ankyrin repeat protein: implications for stability.
H Kaspar Binz1, Andreas Kohl, Andreas Plückthun
1Biochemisches Institut, Universität Zürich, Zürich, Switzerland.
Proteins
|February 24, 2006
Summary
Consensus-designed ankyrin repeat (AR) proteins show high stability. Structural analysis reveals that surface charge distribution, not overall fold, explains stability differences between AR proteins E3_5 and E3_19.
Area of Science:
- Protein Engineering
- Structural Biology
- Biophysics
Background:
- Consensus-designed ankyrin repeat (AR) proteins are engineered for high thermodynamic stability.
- Previous studies linked stability in designed AR proteins like E3_5 to conserved structural motifs and hydrogen-bonding networks.
Purpose of the Study:
- To investigate the structural basis for the significant difference in thermodynamic stability between two highly similar designed AR proteins, E3_5 and E3_19.
- To compare the structural features of E3_19 with E3_5, which exhibits higher stability.
Main Methods:
- X-ray crystallography was used to determine the atomic structure of the designed AR protein E3_19 at 1.9 Å resolution.
- Comparative analysis of structural features, including root-mean-square deviation (RMSD) of C-alpha atoms, surface charge distribution, and exposed hydrophobic residues, between E3_5 and E3_19.
Main Results:
- The crystal structure of E3_19 revealed a regular AR domain fold with characteristic hydrogen-bonding patterns, similar to E3_5.
- Despite 88% sequence identity and virtually identical overall structures (RMSD(Calpha) ≈ 0.7 Å), E3_19 exhibited significantly lower stability (9.6 kcal/mol) compared to E3_5 (14.8 kcal/mol).
- Key differences were identified in the surface properties: E3_19 displayed clusters of charged residues and increased exposure of hydrophobic residues compared to E3_5.
Conclusions:
- The lower thermodynamic stability of the designed AR protein E3_19, compared to E3_5, is not attributed to differences in the overall protein fold or hydrogen-bonding patterns.
- Surface charge distribution and the exposure of hydrophobic residues are critical factors influencing the thermodynamic stability of consensus-designed ankyrin repeat proteins.