Abstract
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
Protein & Peptide Letters
Title: Structural Insights into the Exchange Domain of Sec2p: Expression, Purification,Crystallization, and Preliminary X-Ray Diffraction Data Analysis
Volume: 14 Issue: 3
Author(s): Harindarpal S. Gill
Affiliation:
Keywords: Pseudo-precession plot, Laue symmetry, self-rotation function, non-crystallographic symmetry, coiled-coil, nucleotide exchange factor
Abstract: 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.
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Cite this article as:
Gill S. Harindarpal, Structural Insights into the Exchange Domain of Sec2p: Expression, Purification,Crystallization, and Preliminary X-Ray Diffraction Data Analysis, Protein & Peptide Letters 2007; 14 (3) . https://dx.doi.org/10.2174/092986607780090892
DOI https://dx.doi.org/10.2174/092986607780090892 |
Print ISSN 0929-8665 |
Publisher Name Bentham Science Publisher |
Online ISSN 1875-5305 |
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