Macromolecular X-ray Crystallography Facility

Knowledge of molecular structure is essential for our understanding of biological systems. X-ray crystallography is most widely used to determine the three dimensional structure of proteins, nucleic acids, and large nucleoprotein complexes. It exploits the propensity of X-rays to be scattered by the electron clouds around atoms. The ‘electron density’ can be reconstructed from diffraction patterns obtained from single crystals of the target molecule (shown below). An atomic model is then built into the electron density, which in turn is refined against the data, eventually resulting in an accurate molecular structure in which the coordinates of every atom with respect to every other atom is known precisely.

The Department of Biochemistry and Molecular Biology at Colorado State University has a state-of-the-art macromolecular X-ray crystallography facility. Crystal growth is performed in a climate controlled crystallization room equipped with two bright field/dark field stereomicroscopes, vibration-isolated shelves for long term crystal tray storage, and two 4-60°C high-precision temperature adjustable incubators for crystal growth. We also have the requisite instruments for freezing, mounting, and storing crystals. A Douglas Instruments Oryx crystallization robot is available for screening crystallization conditions.

The X-ray generator consists of a Rigaku RU-H3R rotating anode (target material Cu), which typically operates at 4kW (40 kV, 100 mA) power output. Both X-ray ports are equipped with VariMax X-ray optics, which have characteristics of high flux, excellent spectral purity, uniform beam profile, and adjustable divergence, delivering superior performance. Two imaging plate detectors (R-Axis IV and R-Axis IV++) are installed, which have very low background noise and a large aperture (300mm X 300mm) enabling concurrent collection of strong and weak reflections. Two X-stream 2000 low temperature systems generate their own nitrogen and produce a constant cold nitrogen stream for long periods of time allowing data collection at liquid nitrogen temperatures (-180°C) thus minimizing radiation damage to crystals. Instrumentation has been expanded by a grant from the W. M. Keck foundation, effectively doubling its capacity. The system yields data of exceptional quality and numerous crystal structures have been determined using this system.

The two ports of the X-ray generator are controlled by separate computers which have the latest versions of CrystalClear and d*TREK allowing initial data collection and advanced data processing. These computers have ample data storage capabilities and are also connected to a cluster of Linux workstations. These computers are equipped with all the software required for data reduction (HKL2000, d*TREK and MOSFLM), phase determination (CNS, CCP4 program suite, SOLVE / RESOLVE, BnP), and model building (Coot and Pymol).

The X-ray facilities are open to all appropriately trained CSU faculty, staff and students.