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–T₄ complex has been determined to 3 Å resolution. The monoclinic form of human TTR reported here crystallizes in space group P2₁, with unit-cell parameters a = 76.7 (6), b = 96.7 (8), c = 81.7 (4) Å, β = 106.8 (4)°. The asymmetric unit contains two tetramers of transthyretin related by the non-crystallographic symmetry (NCS) operation of a 90.28° 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^α positions is 0.48 Å. The structure was refined using 15.0–3.0 Å resolution data to R = 22.9% and R_free = 28.9% for reflections F > 0.0σ(F), and R = 19.7% and R_free = 25.8% for reflections F > 3.0σ(F). The intermolecular interactions involve the tips of α-helices and loops around Arg21, Glu61 and Ser100 of all monomers. The electron-density maps revealed residual thyroxine (T₄) 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–T₄ complex. The T₄ 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 β-strands of two monomeric subunits are observed between the apo TTR and T₄-bound tetramer. The averaged distances between Ala108 C^α and its equivalent measured across each binding site are 12.34 Å for the T₄-bound and 10.96 Å 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.