Structures of designed armadillo-repeat proteins show propagation of inter-repeat interface effects

Abstract

The armadillo repeat serves as a scaffold for the development of modular peptide-recognition modules. In order to develop such a system, three crystal structures of designed armadillo-repeat proteins with third-generation N-caps (YIII-Type), four or five internal repeats (M-Type) and second-generation C-caps (AII-Type) were determined at 1.8 A (His-YIIIM4AII), 2.0 A (His-YIIIM5AII) and 1.95 A (YIIIM5AII) resolution and compared with those of variants with thirdgeneration C-caps. All constructs are full consensus designs in which the internal repeats have exactly the same sequence, and hence identical conformations of the internal repeats are expected. The N-cap and internal repeats M1 to M3 are indeed extremely similar, but the comparison reveals structural differences in internal repeats M4 and M5 and the C-cap. These differences are caused by longrange effects of the C-cap, contacting molecules in the crystal, and the intrinsic design of the repeat. Unfortunately, the rigid-body movement of the C-Terminal part impairs the regular arrangement of internal repeats that forms the putative peptide-binding site. The second-generation C-cap improves the packing of buried residues and thereby the stability of the protein. These considerations are useful for future improvements of an armadillo-repeat-based peptiderecognition system.