Crystallization and preliminary crystallographic analysis of cyanide-insensitive alternative oxidase from Trypanosoma brucei brucei

Kido, Yasutoshi, Shiba, Tomoo, Inaoka, Daniel Ken, Sakamoto, Kimitoshi, Nara, Takeshi, Aoki, Takashi, Honma, Teruki, Tanaka, Akiko, Inoue, Masayuki, Matsuoka, Shigeru, Moore, Anthony, Haradaf, Shigeharu and Kita, Kiyoshi (2010) Crystallization and preliminary crystallographic analysis of cyanide-insensitive alternative oxidase from Trypanosoma brucei brucei. Acta Crystallographica Section F, 66 (3). pp. 275-278. ISSN 1744-3091

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Abstract

Cyanide-insensitive alternative oxidase (AOX) is a mitochondrial membrane protein and a non-proton-pumping ubiquinol oxidase that catalyzes the four-electron reduction of dioxygen to water. In the African trypanosomes, trypanosome alternative oxidase (TAO) functions as a cytochrome-independent terminal oxidase that is essential for survival in the mammalian host; hence, the enzyme is considered to be a promising drug target for the treatment of trypanosomiasis. In the present study, recombinant TAO (rTAO) overexpressed in haem-deficient Escherichia coli was purified and crystallized at 293 K by the hanging-drop vapour-diffusion method using polyethylene glycol 400 as a precipitant. X-ray diffraction data were collected at 100 K and were processed to 2.9 resolution with 93.1% completeness and an overall Rmerge of 9.5%. The TAO crystals belonged to the orthorhombic space group I222 or I212121, with unit-cell parameters a = 63.11, b = 136.44, c = 223.06 . Assuming the presence of two rTAO molecules in the asymmetric unit (2 38 kDa), the calculated Matthews coefficient (VM) was 3.2 3 Da 1, which corresponds to a solvent content of 61.0%. This is the first report of a crystal of the membrane-bound diiron proteins, which include AOXs.

Item Type: Article
Schools and Departments: School of Life Sciences > Biochemistry
Subjects: Q Science
Depositing User: EPrints Services
Date Deposited: 06 Feb 2012 18:37
Last Modified: 05 Oct 2012 09:10
URI: http://sro.sussex.ac.uk/id/eprint/17477
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