W0209

Using Protein Crystal Morphology Variations

to Probe Contact Formation Kinetics. Travis Gallagher, Gary Gilliland, Carrie Stover and Wen Pan; Center for Advanced Research in Biotechnology of the University of Maryland Biotechnology Institute and the National Institute of Standards and Technology, 9600 Gudelsky Drive, Rockville, MD 20850, USA

By dissecting the growth of well-characterized crystals into the various intermolecular contacts, it is possible to relate induced changes in crystal growth to chemical effects at the crystal contacts. In one system involving the enzyme subtilisin, we have reported that ionic strength and pH variations cause systematic changes in the shape of the growing crystal. Examination of the crystal contacts in the 1.8 Å resolution X-ray structure leads to the hypothesis that a specific intermolecular hydrogen bond is responsible for the shape effect. This hypothesis has been tested by mutation; in the mutant crystals the shape effect is dramatically reduced, supporting a tentative mechanism involving charge repulsion and cation screening in the nascent contact. This system illustates our method of analyzing contact-formation chemistry by perturbation of the crystal shape, and thus the relative growth rates in various directions, through changes in growth conditions. A second system under study involves a DNA 21-mer whose crystals diffract to 2.5 Å resolution. The DNA crystals belong to space group P6222 or P6422 and grow as hexagonal prisms with an aspect ratio that changes as a function of the divalent cations present.