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Review
. 2009 Aug;19(4):449-57.
doi: 10.1016/j.sbi.2009.04.008. Epub 2009 May 26.

Engineering of recombinant crystallization chaperones

Shohei Koide  1
Affiliations
Review

Engineering of recombinant crystallization chaperones

Shohei Koide. Curr Opin Struct Biol. 2009 Aug.

Abstract

The preparation of diffraction quality crystals remains the major bottleneck in macromolecular X-ray crystallography. A crystallization chaperone is an auxiliary protein, such as fragments of monoclonal antibodies, that binds to and increases the crystallization probability of a target molecule of interest. Such chaperones reduce conformational heterogeneity, mask counterproductive surfaces while extending surfaces predisposed to forming crystal contacts, and provide phasing information. Crystallization chaperones generated using recombinant technologies have emerged as superior alternatives that increase the throughput and eliminate inherent limitations associated with antibody production by animal immunization and the hybridoma technology.

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Figures

Figure 1
Figure 1
The concept of crystallization chaperones. (a) A crystallization chaperone that binds to a specific conformation reduces conformational heterogeneity. (b) A crystallization chaperone can promote crystal lattice formation.
Figure 2
Figure 2
Recent structures determined using crystallization chaperones. (a) β2-adrenergic receptor with Fab (2R4R). (b) Full-length KcsA with a synthetic Fab (3EFF). (c) The P4P6 domain of group I intron with a synthetic Fab. (d) GspD with VHH (3EDJ). (e) Estrogen receptor ligand-binding domain with FN3 (2OCF). (f) AcrB with DARPIN (2J8S).
Figure 3
Figure 3
Crystal packing interactions through exposed edges of β-sheets often observed for β-rich crystallization chaperones. Two different modes of intermolecular β-sheet formation seen in RNaseA-VHH complex structures are shown. (a) 1BZQ (space group, P1) and (b) 2P48 (P3121). β-Strand A of the VHH chaperone is shown as stick models and intermolecular hydrogen bonds are shown as dashed lines. Also see Figure 2d for another example.
Figure 4
Figure 4
Comparisons of the KcsA cytoplasmic domain determined with two different Fab chaperones. (a) The full-length construct with “Fab2”. Only the C-terminal, cytoplasmic domain is shown for KcsA. (b) The truncated construct with “Fab4”. (c) Superposition of the two structures. The RMSD for the common Cα atoms is ∼1 Å.
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