Lessons from high-throughput protein crystallization screening: 10 years of practical experience

Abstract

X-ray crystallography provides the majority of our structural biological knowledge at a molecular level and, in terms of pharmaceutical design, is a valuable tool to accelerate discovery. It is the premier technique in the field, but its usefulness is significantly limited by the need to grow well-diffracting crystals. It is for this reason that high-throughput crystallization has become a key technology that has matured over the past 10 years through the field of structural genomics. Areas covered : The authors describe their experiences in high-throughput crystallization screening in the context of structural genomics and the general biomedical community. They focus on the lessons learnt from the operation of a high-throughput crystallization-screening laboratory, which to date has screened over 12,500 biological macromolecules. They also describe the approaches taken to maximize the success while minimizing the effort. Through this, the authors hope that the reader will gain an insight into the efficient design of a laboratory and protocols to accomplish high-throughput crystallization on a single-, multiuser laboratory or industrial scale. Expert opinion : High-throughput crystallization screening is readily available but, despite the power of the crystallographic technique, getting crystals is still not a solved problem. High-throughput approaches can help when used skillfully; however, they still require human input in the detailed analysis and interpretation of results to be more successful.

Figure 1

A historical perspective of the Hauptman-Woodward high-throughput crystallization screening laboratory illustrating the cumulative and yearly number of macromolecules screened against time and notable events in the laboratory. Detail is given in the text.

Figure 2

High-throughput microbatch-under-oil screening outcomes from our laboratory that show examples of crystal leads for several different proteins. Each image displays a well of 0.9 mm diameter. Some hits are obvious crystals, others need closer examination and are more difficult to discern on the scale of the figure.