NOVEL SYTEMATIC PURIFICATION METHOD DEVELOPMENT TO RAPIDLY OBTAIN HIGH PURITY BIOLOGICAL MOLECULES FOR CRYSTALLOGRAPHY STUDIES. Daryl J. Vanderburgh, Patrick R. Carberry, Michael Meys, PerSeptive Biosystems, Inc. 500 Old Connecticut Path, Framingham, MA 01701

Obtaining adequate quantities of high purity biological molecules that support crystal growth often becomes a frustrating bottleneck that impedes the progress of structural studies. The molecular subjects of these studies have either never been purified before, or existing published purification protocols weren't designed with the purity requirements of crystallography in mind. Crystallographers must either face the tedium of developing new purification methods themselves or get their purified material from collaborators - which can introduce its own set of bottlenecks. Perfusion Chromatographyreg. is a breakthrough technology that allows you to perform chromatographic separations 10 to 100 times faster than with conventional media. With individual run times of 3 to 5 minutes, it now becomes practical to quickly examine all the variables that impact on a separation and to systematically hone in on the best possible purification protocol. It is not uncommon to develop a complete method yielding crystallization quality material in just a few days. Once a method is developed, the fast run times can be equally exploited to generate the necessary quantities of final material for crystallization experiments on an "as-needed" basis - offering the potential for improved experimental flexibility and productivity. Fast run times also help to ensure that purified molecules are recovered in their biologically active form. The principles of Perfusion Chromatography technology will be discussed and some practical examples of its application in structural studies will be shown. The technology has already been eagerly embraced by a number of researchers in the X-ray crystallography field.