Determining the molecular-packing arrangements on protein crystal faces by atomic force microscopy

Previous atomic force microscopy (AFM) studies and periodic bond-chain (PBC) analyses of tetragonal lysozyme crystals have suggested that the (110) face consists of chains of molecules related to one another by 4₃ axes parallel to the crystal face. In this study, high-resolution AFM images of the (110) face were obtained and analyzed in order to verify this prediction. A computer program was employed which constructs the theoretical AFM image corresponding to a specific crystallographic molecular-packing arrangement and AFM tip shape. The packing arrangement and tip shape were varied in order to obtain the maximum possible correlation between experimental and theoretical images. The prediction from PBC analysis of an arrangement involving 4₃ helices was confirmed in this manner, while the alternate arrangement, consisting of molecules related to one another by 2₁ axes, was not observed. However, the surface structure was found to differ significantly even from this crystallographic arrangement. The molecules were found to pack slightly closer about what will become the 4₃ axes within the interior of the crystal, suggesting the occurrence of surface reconstruction or rearrangement on the tetragonal lysozyme (110) face. This study represents a new approach for more precise determination of the molecular-packing arrangements on protein crystal faces employing AFM.