Determining the molecular-growth mechanisms of protein crystal faces by atomic force microscopy.

Abstract

A high-resolution atomic force microscopy (AFM) study has shown that the molecular packing on the tetragonal lysozyme (110) face corresponds to only one of two possible packing arrangements, suggesting that growth layers on this face are of bimolecular height [Li et al. (1999). Acta Cryst. D55, 1023-1035]. Theoretical analyses of the packing also indicated that growth of this face should proceed by the addition of growth units of at least tetramer size, corresponding to the 43 helices in the crystal. In this study, an AFM linescan technique was used to measure the dimensions of individual growth units on protein crystal faces as they were being incorporated into the lattice. Images of individual growth events on the (110) face of tetragonal lysozyme crystals were observed, shown by jump discontinuities in the growth step in the linescan images. The growth-unit dimension in the scanned direction was obtained from these images. A large number of scans in two directions on the (110) face were performed and the distribution of lysozyme growth-unit sizes were obtained. A variety of unit sizes corresponding to 43 helices were shown to participate in the growth process, with the 43 tetramer being the minimum observed size. This technique represents a new application for AFM, allowing time-resolved studies of molecular processes to be carried out.