Characterization of precrystallization aggregation of canavalin by dynamic light scattering

The aggregation processes leading to crystallization and precipitation of canavalin have been investigated by dynamic light scattering (DLS) in photon correlation spectroscopy (PCS) mode. The sizes of aggregates formed under various conditions of pH, salt concentration, and protein concentrations were deduced from the correlation functions generated by the fluctuating intensity of light scattered by the solutions of the protein. Results obtained indicate that the barrier to crystallization of canavalin is the formation of the trimer, a species that has been characterized by x-ray crystallographic studies (McPherson, A. 1980. J. Biol. Chem. 255:10472-10480). The dimensions of the trimer in solution are in good agreement with those obtained both from the crystal (McPherson, A. 1980. J. Biol. Chem. 255:10472-10480) and from a low angle x-ray scattering study in solution (Plietz, P., P. Damaschun, J. J. Müller, and B. Schlener. 1983. FEBS [Fed. Eur. Biochem. Soc.] Lett. 162:43-46). Furthermore, under conditions known to lead to the formation of rhombohedral crystals of canavalin, a limiting size is reached at high concentrations of canavalin. The size measured corresponds to an aggregate of trimers making a unit rhombohedral cell consistent with x-ray crystallographic data (McPherson, A. 1980. J. Biol. Chem. 255:10472-10480). Presumably, such aggregates are the nuclei from which crystal growth proceeds. The present study was undertaken primarily to test the potential of DLS (PCS) as a tool for rapid, routine screening to determine the ultimate fate of protein solutions (i.e., crystallization or amorphous precipitation) at an early stage, therefore eliminating the need for long-term visual observation. Achieving this goal would constitute amajor advance in the practive of protein crystallization. Delays imposed by visual observation would be considerably reduced, and a more systematic approach could be adopted to select experimental conditions.Our findings with canavalin demonstrate that DLS(PCS) is, indeed, a selective and sensitive probe of precrystallization conditions. Other advantages of this technique include the facts that it is noninvasive, nondestructive,universal, and does not require calibration.