Overproduction, purification, crystallization and preliminary X-ray diffraction analysis of Trypanosoma brucei gambiense glycerol kinase

Balogun, Emmanuel Oluwadare;Inaoka, Daniel Ken;Kido, Yasutoshi;Shiba, Tomoo;Kita, Kiyoshi;et.al.
(2010) Acta Crystallographica Section F-structural Biology And Crystallization Communications — Vol. 66, n° Pt 3, p. 304-308 (2010)

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Authors
  • Balogun, Emmanuel Oluwadare
    Author
  • Inaoka, Daniel Ken
    Author
  • Kido, Yasutoshi
    Author
  • Shiba, Tomoo
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  • Michels, PaulusUCLouvain
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  • Kita, Kiyoshi
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Abstract
In the bloodstream forms of human trypanosomes, glycerol kinase (GK; EC 2.7.1.30) is one of the nine glycosomally compartmentalized enzymes that are essential for energy metabolism. In this study, a recombinant Trypanosoma brucei gambiense GK (rTbgGK) with an N-terminal cleavable His(6) tag was overexpressed, purified to homogeneity and crystallized by the sitting-drop vapour-diffusion method using PEG 400 as a precipitant. A complete X-ray diffraction data set to 2.75 angstrom resolution indicated that the crystals belonged to the orthorhombic space group P2(1)2(1)2(1), with unit-cell parameters a = 63.84, b = 121.50, c = 154.59 angstrom. The presence of two rTbgGK molecules in the asymmetric unit gives a Matthews coefficient (V-M) of 2.5 angstrom(3) Da(-1), corresponding to 50% solvent content.
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Balogun, E. O., Inaoka, D. K., Kido, Y., Shiba, T., Nara, T., Aoki, T., Honma, T., Tanaka, A., Inoue, M., Matsuoka, S., Michels, P., Harada, S., & Kita, K. (2010). Overproduction, purification, crystallization and preliminary X-ray diffraction analysis of Trypanosoma brucei gambiense glycerol kinase. Acta Crystallographica Section F-structural Biology And Crystallization Communications, 66(Pt 3), 304-308. https://doi.org/10.1107/S1744309110000369 (Original work published 2010)