Structure of a new polymorphic monoclinic form of human transthyretin at 3 Å resolution reveals a mixed complex between unliganded and T4-bound tetramers of TTR

The crystal structure of a new polymorphic form of human transthyretin (hTTR) with a lattice containing a unique assembly of apo hTTR and TTR-T-4 complex has been determined to 3 Angstrom resolution. The monoclinic form of human TTR reported here crystallizes in space group P2(1), with unit-cell parameters a = 76.7 (6), b = 96.7 (8), c = 81.7 (4) Angstrom, beta = 106.8 (4)degrees. The asymmetric unit contains two tetramers of transthyretin related by the non-crystallographic symmetry (NCS) operation of a 90.28 degrees rotation between two hTTR molecules around an axis close to crystallographic z. The r.m.s. difference between the two tetramers calculated from their C-alpha positions is 0.48 Angstrom. The structure was refined using 15.0-3.0 Angstrom resolution data to R = 22.9% and R-free = 28.9% for reflections F>0.0 sigma (F), and R = 19.7% and R-free = 25.8% for reflections F >3.0 sigma (F). The intermolecular interactions involve the tips of alpha -helices and loops around Arg21, Glu61 and Ser100 of all monomers. The electron-density maps revealed residual thyroxine (T-4) bound in only one of the two unique tetrameric TTR molecules, with an occupancy of 53%, while the second tetramer is unliganded. One thyroxine ligand is bound in a way similar to the orientations described for the orthorhombic form of the hTTR-T-4 complex. The T-4 bound in the second site is positioned similar to 3',5'-dinitro-N-acetyl-L-thyronine in its hTTR complex. Differences in the size of the central channel defined by the D, A, G and H beta -strands of two monomeric subunits are observed between the apo TTR and T-4-bound tetramer. The averaged distances between Ala108 C-alpha and its equivalent measured across each binding site are 12.34 Angstrom for the T-4-bound and 10.96 Angstrom for the unliganded TTR tetramer, respectively. The observed differences might reflect the mechanics of the ligand binding in the channel and possibly explain the observed negative cooperativity effect for ligand binding.