Crystallizing proteins on the basis of their precipitation diagram determined using a microfluidic formulator.
Crystallization of proteins from a purified protein solution remains a bottleneck in the structure determination pipeline. In this paper the crystallization problem is addressed using a microfluidic device capable of determining detailed protein precipitation diagrams using less than 10 microL of protein sample. Based on the experimentally determined protein phase behavior, a crystallization screen can be designed to accommodate the physical chemistry of the particular protein target. Such a tailor-made crystallization screen has a high probability of yielding crystallization hits. The approach is applied to two different proteins: the calcium pump (SERCA), an eukaryotic integral membrane protein, and UMP kinase, a prokaryotic soluble kinase. Protein phase behavior is mapped for both proteins and tailor-made crystallization screens are designed for the two proteins resulting in about 50% crystallization probability per experiment. This illustrates the power of using microfluidic devices for detailed characterization of protein phase behavior prior to crystallization trials.
Sommer
Morten O A
MO
Centre for Crystallographic Studies, Department of Chemistry, University of Copenhagen, Denmark. morten@ccs.ki.ku.dk
Larsen
Sine
S
D000818:Animals; D000252:Calcium-Transporting ATPases / Q000737:chemistry*; D011232:Chemical Precipitation; D003460:Crystallization / Q000379:methods*; D006863:Hydrogen-Ion Concentration; D044085:Microfluidics / Q000295:instrumentation*; D009703:Nucleoside-Phosphate Kinase / Q000737:chemistry*; D011817:Rabbits
C023276:uridine monophosphate kinase; D009703:Nucleoside-Phosphate Kinase; D000252:Calcium-Transporting ATPases