A role for native lipids in the stabilization and two dimensional crystallization of the Escherichia coli NADH ubiquinone oxidoreductase (complex I) Journal Article


Author(s): Sazanov, Leonid A; Carroll, Joe D; Holt, Peter J; Toime, Laurence J; Fearnley, Ian M
Article Title: A role for native lipids in the stabilization and two dimensional crystallization of the Escherichia coli NADH ubiquinone oxidoreductase (complex I)
Affiliation
Abstract: NADH-ubiquinone oxidoreductase (complex I or NDH-1) was purified from the BL21 strain of Escherichia coli using an improved procedure. The complex was effectively stabilized by addition of divalent cations and lipids, making the preparation suitable for structural studies. The ubiquinone reductase activity of the enzyme was fully restored by addition of native E. coli lipids. Two different two-dimensional crystal forms, with p2 and p3 symmetry, were obtained using lipids containing native E. coli extracts. Analysis of the crystals showed that they are formed by fully intact complex I in an L-shaped conformation. Activity assays and single particle analysis indicated that complex I maintains this structure in detergent solution and does not adopt a different conformation in the active state. Thus, we provide the first experimental evidence that complex I from E. coli has an L-shape in a lipid bilayer and confirm that this is also the case for the active enzyme in solution. This suggests strongly that bacterial complex I exists in an L-shaped conformation in vivo. Our results also indicate that native lipids play an important role in the activation, stabilization and, as a consequence, crystallization of purified complex I from E. coli.
Keywords: Escherichia coli; Enzymes; Crystallization; Crystals; Detergents; Solutions Two-dimensional crystallization Lipids
Journal Title: Journal of Biological Chemistry
Volume: 278
Issue 21
ISSN: 1083-351X
Publisher: American Society for Biochemistry and Molecular Biology  
Date Published: 2003-05-23
Start Page: 19483
End Page: 19491
DOI: 10.1074/jbc.M208959200
Open access: no