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Protein Sci. 2010 May;19(5):901-13. doi: 10.1002/pro.368.

A synergistic approach to protein crystallization: combination of a fixed-arm carrier with surface entropy reduction.

Author information

1
Laboratory of Structural Biology, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina 27709, USA.

Abstract

Protein crystallographers are often confronted with recalcitrant proteins not readily crystallizable, or which crystallize in problematic forms. A variety of techniques have been used to surmount such obstacles: crystallization using carrier proteins or antibody complexes, chemical modification, surface entropy reduction, proteolytic digestion, and additive screening. Here we present a synergistic approach for successful crystallization of proteins that do not form diffraction quality crystals using conventional methods. This approach combines favorable aspects of carrier-driven crystallization with surface entropy reduction. We have generated a series of maltose binding protein (MBP) fusion constructs containing different surface mutations designed to reduce surface entropy and encourage crystal lattice formation. The MBP advantageously increases protein expression and solubility, and provides a streamlined purification protocol. Using this technique, we have successfully solved the structures of three unrelated proteins that were previously unattainable. This crystallization technique represents a valuable rescue strategy for protein structure solution when conventional methods fail.

PMID:
20196072
PMCID:
PMC2868234
DOI:
10.1002/pro.368
[Indexed for MEDLINE]
Free PMC Article

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