MOLECULAR-BASIS FOR IONIC-STRENGTH DEPENDENCE AND CRYSTAL MORPHOLOGY IN 2-DIMENSIONAL STREPTAVIDIN CRYSTALLIZATION

The two-dimensional crystallization of streptavidin at functionalized lipid interfaces is one of the best studied model systems for investig ating molecular self-assembly processes. This system also provides an opportunity to elucidate the relationship between protein-protein mole cular recognition, crystallization solution conditions, and crystal pr operties such as coherence, space group symmetry, and morphology. A be tter understanding of these relationships may aid in the design of rat ional strategies for promoting high-quality protein crystallization an d for controlling protein assembly at interfaces in the biomaterials a nd nanotechnology fields. Here we show that two-dimensional streptavid in crystallization is kinetically controlled and that formation of a s ingle electrostatic interaction at the crystal contact interfaces is a key energetic determinant of the kinetic barriers controlling crystal morphology. Our results also demonstrate that this electrostatic inte raction at the streptavidin protein-protein interfaces is responsible for the ionic strength dependence of streptavidin crystallization. Mol ecular modeling studies of the wild-type crystal that displays C222 sy mmetry suggested that the side-chain amines of lysine 132 from adjacen t proteins interact with each other across the dyad-related crystal co ntacts. Leucine was substituted at this position (K132L) to remove the need for bridging counterions. Unlike wild-type streptavidin, the K13 2L mutant crystallizes with rectangular morphology on buffer or on a p ure water subphase and analysis of the electron micrographs demonstrat es that the crystal retains C222 symmetry in the presence or absence o f salt. The kinetic barriers associated with formation of this electro static interaction thus underlie the wild-type butterfly crystal morph ology.