Crystallizing proteins on the basis of their precipitation diagram determined using a microfluidic formulator

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Abstract

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 μL 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.
Original languageEnglish
JournalJournal of Synchrotron Radiation
Volume12
Issue number6
Pages (from-to)779-785
Number of pages7
ISSN0909-0495
DOIs
Publication statusPublished - 2005
Externally publishedYes

Keywords

  • Microfluidics
  • Crystallization
  • Membrane proteins
  • Kinases
  • Phase behavior
  • Physical characterization

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