Overproduction, purification, crystallization and preliminary X-ray diffraction analysis of Trypanosoma brucei gambiense glycerol kinase.
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 A 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 A. The presence of two rTbgGK molecules in the asymmetric unit gives a Matthews coefficient (V(M)) of 2.5 A(3) Da(-1), corresponding to 50% solvent content.
Balogun
Emmanuel Oluwadare
EO
Department of Biomedical Chemistry, Graduate School of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
Inaoka
Daniel Ken
DK
Kido
Yasutoshi
Y
Shiba
Tomoo
T
Nara
Takeshi
T
Aoki
Takashi
T
Honma
Teruki
T
Tanaka
Akiko
A
Inoue
Masayuki
M
Matsuoka
Shigeru
S
Michels
Paul A M
PA
Harada
Shigeharu
S
Kita
Kiyoshi
K
D000818:Animals; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D003460:Crystallization; D018360:Crystallography, X-Ray; D005991:Glycerol Kinase / Q000737:chemistry* / Q000302:isolation & purification; D014347:Trypanosoma brucei gambiense / Q000201:enzymology*
D005991:Glycerol Kinase