Comparison of the decameric structure of peroxiredoxin-II by transmission electron microscopy and X-ray crystallography

The decameric human erythrocyte protein torin is identical to the thiol-spe cific antioxidant protein-II (TSA-II), also termed peroxiredoxin-II (Prx-II ). Single particle analysis from electron micrographs of Prx-II molecules h omogeneously orientated across holes in the presence of a thin film of ammo nium molybdate and trehalose has facilitated the production of a greater th an or equal to 20 Angstrom 3-D reconstruction by angular reconstitution tha t emphasises the D5 symmetry of the ring-like decamer. The X-ray structure for Prx-II was fitted into the transmission electron microscopic reconstruc tion by molecular replacement. The surface-rendered transmission electron m icroscopy (TEM) reconstruction correlates well with the solvent-excluded su rface of the X-ray structure of the Prx-II molecule. This provides confirma tion that transmission electron microscopy of negatively stained specimens, despite limited resolution, has the potential to reveal a valid representa tion of surface features of protein molecules. 2-D crystallisation of the P rx-II protein on mica as part of a TEM study resulted in the formation of a p2 crystal form with parallel linear arrays of stacked rings. This latter 2-D form correlates well with that observed from the 2.7 Angstrom X-ray str ucture of Prx-II solved from a new orthorhombic 3-D crystal form. (C) 2001 Elsevier Science B.V. All rights reserved.