title: Molecular symmetry-constrained systematic search approach to structure solution of the coiled-coil SRGAP2 F-BARx domain.
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.
authors.affiliation: The Mina and Everard Goodman Faculty of Life Sciences, Bar-Ilan University, Ramat Gan 52900, Israel.
authors.forename: Michael
authors.initials: M
authors.lastname: Sporny
authors.affiliation: The Mina and Everard Goodman Faculty of Life Sciences, Bar-Ilan University, Ramat Gan 52900, Israel.
authors.forename: Julia
authors.initials: J
authors.lastname: Guez-Haddad
authors.affiliation: STFC Rutherford Appleton Laboratory, Didcot, England.
authors.forename: David G
authors.initials: DG
authors.lastname: Waterman
authors.affiliation: Biosciences, University of Exeter, Exeter, England.
authors.forename: Michail N
authors.initials: MN
authors.lastname: Isupov
authors.affiliation: The Mina and Everard Goodman Faculty of Life Sciences, Bar-Ilan University, Ramat Gan 52900, Israel.
authors.forename: Yarden
authors.initials: Y
authors.lastname: Opatowsky
mesh_terms: D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D018360:Crystallography, X-Ray; D020690:GTPase-Activating Proteins / Q000737:chemistry*; D006801:Humans; D008958:Models, Molecular; D000072756:Protein Conformation, alpha-Helical; D000072417:Protein Domains
chemical_list: D020690:GTPase-Activating Proteins; C501995:SRGAP2 protein, human
keywords: F-BAR; SRGAP2; coiled coil; exhaustive search; molecular replacement