Technological advances have increased the value of the laboratory diffractometer for protein crystallography. To find out how different labs benefit from the latest equipment, DECTRIS sent two adventurers on a tour along the East Coast of the United States of America. There, three laboratories with EIGER R 4M detectors are located within a short driving distance from one another.
Martin Safo, Professor in the Department of Medicinal Chemistry at VCU, hasn't collected at a synchrotron in a while. He works on small molecules that increase the affinity of hemoglobin for oxygen, a potential approach to treating sickle cell disease. The key is to characterize synthesized compounds and ligand-bound targets quickly and accurately. His rotating anode diffractometer with an EIGER R 4M detector works wonderfully.
Faik Musayev, the manager of the X-ray facility, routinely collects data on small-molecule crystals to a resolution of 0.83 Å despite working with copper radiation. The large active area of the detector and the two-theta arm of the diffractometer make that possible. On the same system, without any configuration changes, he collects data to better than 2 Å resolution from ligand-bound protein crystals. This makes identification of the ligand and characterization of its binding mode possible. "We can get all the information we need," says Musayev. Medicinal chemists in the departments then improve the small molecule based on this information - without being held back by synchrotron beamtime schedules.
In one year of working with their new system, Safo and Musayev have already deposited four structures in the PDB (6BNR, 6BWP, 6BWU, 6DI4). They have characterized two classes of allosteric effectors for the treatment of sickle cell disease (Pagare 2018 and Nakagawa 2018) and identified promising lead candidates for in vivo pharmacologic studies and further structural optimization.
Virginia Commonwealth University X-ray facility manager Faik Musayev and Prof Martin Safo
Fred Dyda at the National Institute of Diabetes and Digestive and Kidney Diseases of the NIH has a long-standing interest in protein DNA interactions. He recently upgraded his bespoke diffractometer with an EIGER R 4M detector. The diffractometer is primarily used for screening crystals destined to the synchrotron. With the noise-free EIGER, even the weakest crystals can be characterized properly before data collection. Dyda can optimize cryo-protection conditions iteratively within a short time. He doesn’t have to send crystals to the synchrotron untested in the hope that their diffraction will be sufficient.
Well-diffracting crystals can be measured right away, with results comparable to what can be achieved at the synchrotron and often driving further experiments. This synergy between structural work and biochemistry is one of the key advantages of a laboratory diffractometer. "You can have your answer without waiting for the next synchrotron slot," says Dyda.
The diffractometer is also a valuable teaching tool. With synchrotron data collected remotely and processed automatically for the most part, students are at the risk of seeing crystallography as something virtual. In a laboratory, they see it's real. Dyda's diffractometer has an inverted phi goniometer that makes mounting crystals easy even for beginners. The detector gives results quickly and of the same kind and quality the students will later encounter at the synchrotron. Students can explore and learn without the pressure of a tight beamtime schedule where high throughput is more important than comprehension.
National Institutes of Health research associate Ivana Grabundzija
Alexander Wlodawer at the NCI in Frederick studies the enzymatic mechanism of L-asparaginase, an enzyme used as a drug against childhood leukemia. His work involves the soaking of putative substrates into protein crystals. Depending on whether and where the reaction intermediates are bound, new conditions are tested and the cycle repeats until the mechanism is revealed. "Having excellent data collection equipment in the lab is crucial when each experiment yields information on what should follow. This practice is not feasible on a synchrotron where we have access only every two to three weeks," says Wlodawer.
On the dual-port rotating anode diffractometer with two EIGER R 4M detectors, Jacek Lubkowski, the staff scientist working on the project, could collect a dozen full datasets a week. Most of the time, biochemistry and crystallization slow him down a bit. Even so, this efficiency would not be possible without a well-equipped diffraction laboratory.
National Cancer Institute X-ray laboratory manager Mi Li
A state-of-the-art laboratory diffractometer makes work easier and research groups more efficient. The key is the immediate feedback you get about your work: Do your crystals diffract sufficiently? Did you soak the right compound? Is the way you cryo-protect your crystal detrimental to diffraction? Does the cryo-structure represent the situation at room temperature? If answering these questions entails a wait for the next synchrotron shift, you're holding yourself back.
Three very different laboratories have upgraded their diffractometers to make exciting science possible and improve their productivity. Yours could be the next. To embark on a journey to immediate results and better data, you don't have to hit the road yourself. Contact our OEM partners Rigaku, marXperts or STOE now and enquire about a diffractometer with an EIGER detector.