Diffraction from Microvolume Topaz Microprocessors

DESCRIPTION (provided by applicant): We propose to develop an efficient and rapid X-ray diffraction screen for protein crystals that will accelerate structure determination of proteins that are difficult to express and purify. Determination of protein structures at atomic resolution has allowed major advances in our fundamental understanding of biology and has provided a basis for rational drug design. However, a major obstacle to determining protein structures by X-ray crystallography has been in the initial step of identifying X-ray diffracting protein crystals. Current methods that identify protein crystals primarily by morphology followed by diffraction are costly in terms of time and material. We will modify the Fluidigm Topaz crystallization chip so that protein crystals can be screened in situ for X-ray diffraction at a synchrotron. We will optimize the material composition of the chip to maximize X-ray transparency while maintaining the functional ability to crystallize proteins. We will then develop the parameters to differentiate protein and contaminating salt crystals that are as small as 10 microns. We will collaborate with multiple structural biologists to utilize the diffraction screen with a wide variety of transmembrane, signaling, and enzymatic proteins. We anticipate that the diffraction screen that we will develop will be easily adapted to other synchrotron facilities and will accelerate crystallographic studies of proteins with scientific and medical importance.