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

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• 作者：Michael Sporny ; Julia Guez-Haddad ; David G. Waterman ; Michail N. Isupov and Yarden Opatowsky
• 关键词：exhaustive search ; SRGAP2 ; F-BAR ; coiled coil ; molecular replacement
• 刊名：Acta Crystallographica Section D
• 出版年：2016
• 出版时间：December 2016
• 年：2016
• 卷：72
• 期：12
• 页码：1241-1253
• 全文大小：1574K
• ISSN：1399-0047

文摘

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.