Reconstitution of mitochondrial ATP synthase into lipid bilayers for structural analysis

Arechaga, Ignacio; Fotiadis, Dimitrios (2007). Reconstitution of mitochondrial ATP synthase into lipid bilayers for structural analysis. Journal of structural biology, 160(3), pp. 287-94. San Diego, Calif.: Elsevier 10.1016/j.jsb.2007.09.007

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Mitochondrial F(1)F(o)-ATP synthase is a molecular motor that couples the energy generated by oxidative metabolism to the synthesis of ATP. Direct visualization of the rotary action of the bacterial ATP synthase has been well characterized. However, direct observation of rotation of the mitochondrial enzyme has not been reported yet. Here, we describe two methods to reconstitute mitochondrial F(1)F(o)-ATP synthase into lipid bilayers suitable for structure analysis by electron and atomic force microscopy (AFM). Proteoliposomes densely packed with bovine heart mitochondria F(1)F(o)-ATP synthase were obtained upon detergent removal from ternary mixtures (lipid, detergent and protein). Two-dimensional crystals of recombinant hexahistidine-tagged yeast F(1)F(o)-ATP synthase were grown using the supported monolayer technique. Because the hexahistidine-tag is located at the F(1) catalytic subcomplex, ATP synthases were oriented unidirectionally in such two-dimensional crystals, exposing F(1) to the lipid monolayer and the F(o) membrane region to the bulk solution. This configuration opens a new avenue for the determination of the c-ring stoichiometry of unknown hexahistidine-tagged ATP synthases and the organization of the membrane intrinsic subunits within F(o) by electron microscopy and AFM.

Item Type:

Journal Article (Original Article)

Division/Institute:

04 Faculty of Medicine > Pre-clinic Human Medicine > Institute of Biochemistry and Molecular Medicine

UniBE Contributor:

Fotiadis, Dimitrios José

ISSN:

1047-8477

ISBN:

17959389

Publisher:

Elsevier

Language:

English

Submitter:

Factscience Import

Date Deposited:

04 Oct 2013 14:53

Last Modified:

04 May 2014 23:15

Publisher DOI:

10.1016/j.jsb.2007.09.007

PubMed ID:

17959389

Web of Science ID:

000251473500004

URI:

https://boris.unibe.ch/id/eprint/22700 (FactScience: 36075)

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