Structural Insights into the Exchange Domain of Sec2p: Expression, Purification,Crystallization, and Preliminary X-Ray Diffraction Data Analysis

Author(s): Harindarpal S. Gill.

Journal Name:Protein & Peptide Letters

Volume 14 , Issue 3 , 2007


Sec2p is an essential yeast gene and is part of the cell polarization process that leads to budding. The Nterminal domain of sec2p (Sec2pN) the guanine-nucleotide exchange factor for sec4p has been expressed in Escherichia coli, purified, and crystallized. Crystals belong to the space group P21 with unit cell dimensions 178.1 x 98.4 x 180.0 Å,β = 91.7°, and diffract synchrotron-generated X-rays to better than 3.6 Å resolution. Pseudo-precession plots reveal a Laue symmetry of 2/m, corresponding to the aforementioned space group, and unusual weak diffraction in the ∼5-7 Å resolution range. The Matthews number calculations for a typical crystal density suggest a range of 28 to 64 molecules per asymmetric unit. Self-rotation and native Patterson calculations demonstrate a pure helical array of protein subunits. Based on the X-ray diffraction data analysis and amino-acid sequence alignments, the paper presents a hypothetical model of the exchange domain of sec2p as a pair of coiled-coil helices that binds to sec4p and facilitates nucleotide disassociation.

Keywords: Pseudo-precession plot, Laue symmetry, self-rotation function, non-crystallographic symmetry, coiled-coil, nucleotide exchange factor

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Article Details

Year: 2007
Page: [253 - 258]
Pages: 6
DOI: 10.2174/092986607780090892
Price: $58

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