Structural Insights into the Exchange Domain of Sec2p: Expression, Purification,Crystallization, and Preliminary X-Ray Diffraction Data Analysis
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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Document Type: Research Article
Affiliations: Department of Cellular and Molecular Physiology, Yale University School of Medicine, 333 Cedar Street, New Haven, CT 06520, USA.
Publication date: 2007-03-01
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- Protein & Peptide Letters publishes short papers in all important aspects of protein and peptide research, including structural studies, recombinant expression, function, synthesis, enzymology, immunology, molecular modeling, drug design etc. Manuscripts must have a significant element of novelty, timeliness and urgency that merit rapid publication. Reports of crystallisation, and preliminary structure determinations of biologically important proteins are acceptable. Purely theoretical papers are also acceptable provided they provide new insight into the principles of protein/peptide structure and function.