Two-dimensional crystallization of Escherichia coli lactose permease

Zhuang, J. and Prive, G. G. and Werner, G. E. and Ringler, P. and Kaback, H. R. and Engel, A.. (1999) Two-dimensional crystallization of Escherichia coli lactose permease. Journal of Structural Biology, Vol. 125, H. 1. pp. 63-75.

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Official URL: http://edoc.unibas.ch/dok/A5257680

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A chimeric protein consisting of lactose permease with cytochrome b562 in the middle cytoplasmic loop and six His residues at the C terminus (LacY/L6cytb562/417H6 or "red permease") was overexpressed in Escherichia coli and isolated by nickel affinity chromatography after solubilization with dodecyl-beta,d-maltopyranoside. Red permease was then reconstituted in the presence of phospholipids, yielding densely packed vesicles and well-ordered two-dimensional (2D) crystals as shown by electron microscopy of negatively stained specimens. Single-particle analysis of 16 383 protein particles in densely packed vesicles reveals a 5.4-nm-long trapeziform protein of 4.1 to 5.1 nm width, with a central stain-filled indentation. Depending on reconstitution conditions, trigonal and rectangular crystallographic packing arrangements of these elongated particles assembled into trimers are observed. The best ordered 2D crystals exhibit a rectangular unit cell, of dimensions a = 9.9 nm, b = 17.4 nm, that houses two trimeric complexes. Projection maps calculated to a resolution of 2 nm show that these crystals consist of two layers.
Faculties and Departments:05 Faculty of Science > Departement Biozentrum > Former Organization Units Biozentrum > Structural Biology (Engel)
UniBasel Contributors:Engel, Andreas H
Item Type:Article, refereed
Article Subtype:Research Article
Publisher:Academic Press
Note:Publication type according to Uni Basel Research Database: Journal article
Last Modified:22 Mar 2012 14:20
Deposited On:22 Mar 2012 13:18

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