ELECTRONS AND X-RAYS WORKING TOGETHER TO VISUALIZE ANTIBODY-RHINOVIRUS INTERACTIONS. Timothy S. Baker, Thomas J. Smith, and Norman H. Olson, Department of Biological Sciences, Purdue University, West Lafayette IN 47907-1392.
Examinations of detailed interactions that occur among macromolecules within large complexes often require or benefit from combined structural information obtained from complementary techniques. Cryo-electron microscopy (cryoEM) and three-dimensional (3D) image reconstruction provide a low resolution envelope or framework for constructing a "pseudo" atomic model of the complex from high resolution structures of the components.
Our 25Å resolution, 3D reconstruction1 of a complex between intact rhinovirus serotype 14 (HRV14) and the Fab fragment of a neutralizing monoclonal antibody (Fab17-IA) was used in this manner to dock, as rigid bodies, the separately determined X-ray structures of the virus2 and Fab fragment3. This model was then used to initiate phasing to 8Å of recently obtained X-ray data from frozen, single crystals4 of the HRV14/Fab17-IA complex.
Preliminary analysis of the X-ray structrure of the complex, after phase extension to 4Å, shows that the Fab CDR3 loop of the heavy chain adjusts its conformation to give a tighter Fab-virus interaction. In addition, the variable domain of the Fab is rotated slightly relative to the starting model in an orientation that fits the cryoEM reconstruction even better than the original model. In hindsight, the decision to dock atomic models as rigid bodies (thereby generating density overlaps) rather than allowing for conformational flexibility in the Fab or virus structures, reduced the overall quality of the initial model. Despite these errors, the initial model led to successful phase extension of the X-ray crystallographic data.
References
1. Smith, T. J., et al. (1993) J. Virol. 67:1148-1158.
2. Rossmann, M. G., et al. (1985) Nature (London) 317:145-153.
3. Liu, H., et al. (1994) J. Mol. Biol. 240:127-137.
4. Smith, T. J.and E. S. Chase (1992) Virology 191:600-606.