CRYSTAL-STRUCTURES OF CHAIN-FOLDED ANTIPARALLEL BETA-SHEET ASSEMBLIESFROM SEQUENCE-DESIGNED PERIODIC POLYPEPTIDES

The crystal structures and textures of a family of sequence-designed p eriodic polypeptides were investigated and analyzed using X-ray diffra ction, vibrational spectroscopy, and cross-polarization magic angle sp inning C-13 nuclear magnetic resonance. The repetitive amino acid sequ ences are described by -[(AG)(x)EG]-, with integer x from 3 to 6. Thes e macromolecules were prepared via bacterial expression of artificial genes and are monodisperse. Crystalline samples were obtained, and the interpretation of the X-ray diffraction results was aided by the gene ration of computer-simulated X-ray diffraction patterns. This allowed direct comparisons to be made with the observed texture-oriented X-ray diffraction photographs. All diffraction and spectroscopic evidence s upports an antiparallel tap) beta-sheet structure, and all structures index on orthorhombic sublattices similar to those reported for Bombyx mori silk fibroin and poly(L-alanylglycine). The unit cell parameters for poly(AG)(3)EG, for example, are a = 0.948 nm (hydrogen-bond direc tion), b = 1.060 nm (ap beta-sheet stacking direction), and c = 0.695 nm (chain direction). Selective line broadening is observed for wide-a ngle diffraction signals with l not equal 0 (for the 211 in particular ) and gives an estimated crystal size of <4 nm in the chain direction. This, coupled with the appearance of a low-angle particle interferenc e peak at 3.6 nm, indicates a crystal size over an order of magnitude less than the chain length and suggests an adjacent reentry chain-fold ed lamellar structure incorporating the ap beta-sheet architecture. A structure with polar ap beta-sheets and gamma-turns, stacking with the hydrophobic methyl groups of the alanyl residues in contact, is selec ted by X-ray structure refinement to give the best match with the expe rimental data. The pattern of crystallization behavior of the poly(AG) ,EG family is consistent with the folding periodicity being in-phase w ith the amino acid sequence so that the glutamic acid residues are con fined to the lamellar surfaces.