RESUMEN

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.

Asunto(s)

Proteínas Represoras/química , Proteínas Represoras/metabolismo , Proteínas Virales/química , Proteínas Virales/metabolismo , Bacteriófago P22/química , Bacteriófago P22/genética , Cristalografía por Rayos X , ADN Viral/genética , ADN Viral/metabolismo , Cinética , Modelos Moleculares , Unión Proteica , Pliegue de Proteína , Estructura Secundaria de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Proteínas Represoras/genética , Proteínas Virales/genética , Proteínas Reguladoras y Accesorias Virales