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Acta Crystallogr D Struct Biol. 2016 Dec 1;72(Pt 12):1241-1253. Epub 2016 Nov 29.

Molecular symmetry-constrained systematic search approach to structure solution of the coiled-coil SRGAP2 F-BARx domain.

Author information

1
The Mina and Everard Goodman Faculty of Life Sciences, Bar-Ilan University, Ramat Gan 52900, Israel.
2
STFC Rutherford Appleton Laboratory, Didcot, England.
3
Biosciences, University of Exeter, Exeter, England.

Abstract

SRGAP2 (Slit-Robo GTPase-activating protein 2) is a cytoplasmic protein found to be involved in neuronal branching, restriction of neuronal migration and restriction of the length and density of dendritic postsynaptic spines. The extended F-BAR (F-BARx) domain of SRGAP2 generates membrane protrusions when expressed in COS-7 cells, while most F-BARs induce the opposite effect: membrane invaginations. As a first step to understand this discrepancy, the F-BARx domain of SRGAP2 was isolated and crystallized after co-expression with the carboxy domains of the protein. Diffraction data were collected from two significantly non-isomorphous crystals in the same monoclinic C2 space group. A correct molecular-replacment solution was obtained by applying a molecular symmetry-constrained systematic search approach that took advantage of the conserved biological symmetry of the F-BAR domains. It is shown that similar approaches can solve other F-BAR structures that were previously determined by experimental phasing. Diffraction data were reprocessed with a high-resolution cutoff of 2.2 Å, chosen using less strict statistical criteria. This has improved the outcome of multi-crystal averaging and other density-modification procedures.

KEYWORDS:

F-BAR; SRGAP2; coiled coil; exhaustive search; molecular replacement

PMID:
27917825
DOI:
10.1107/S2059798316016697
[Indexed for MEDLINE]

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