Consolidating critical binding determinants by noncyclic rearrangement of protein secondary structure.

Publication Type:

Journal Article


Proc Natl Acad Sci U S A, Volume 102, Issue 7, p.2305-9 (2005)


Bacteriophage P22, Crystallography, X-Ray, DNA, Viral, Kinetics, Models, Molecular, Protein Binding, Protein Folding, Protein Structure, Secondary, Recombinant Proteins, Repressor Proteins, Viral Proteins, Viral Regulatory and Accessory Proteins


<p>We designed a single-chain variant of the Arc repressor homodimer in which the beta strands that contact operator DNA are connected by a hairpin turn and the alpha helices that form the tetrahelical scaffold of the dimer are attached by a short linker. The designed protein represents a noncyclic permutation of secondary structural elements in another single-chain Arc molecule (Arc-L1-Arc), in which the two subunits are fused by a single linker. The permuted protein binds operator DNA with nanomolar affinity, refolds on the sub-millisecond time scale, and is as stable as Arc-L1-Arc. The crystal structure of the permuted protein reveals an essentially wild-type fold, demonstrating that crucial folding information is not encoded in the wild-type order of secondary structure. Noncyclic rearrangement of secondary structure may allow grouping of critical active-site residues in other proteins and could be a useful tool for protein design and minimization.</p>