Expression, purification and preliminary crystallographic studies of human ketohexokinase.
Ketohexokinase (KHK; E.C. 2.7.1.3) catalyses the (reversible) phosphorylation of fructose to fructose-1-phosphate. KHK is the first enzyme in a specialized catabolic pathway metabolizing dietary fructose to the glycolytic intermediate glyceraldehyde-3-phosphate. Mutations inactivating KHK underlie the metabolic disorder essential fructosuria. The primary structure of KHK shows no significant homology to other mammalian hexokinases. It is most similar to prokaryotic ribokinases, but catalyses a distinct phosphorylation reaction. Recombinant human KHK has been crystallized in the orthorhombic form (space group P2(1)2(1)2 or P2(1)2(1)2(1)). Single crystals of this polymorph suitable for X-ray diffraction have been obtained by vapour diffusion using 2-propanol and MPD as precipitants (pH 7.5). The crystals have unit-cell parameters a = 93.4, b = 121.5, c = 108.4 A. Diffraction data were collected to 4.3 A resolution. The asymmetric unit contains four protein molecules.
Kozak
M
M
Department of Macromolecular Physics, Faculty of Physics, A. Mickiewicz University, Umultowska 85, 61-614 Poznañ, Poland. mkozak@main.amu.edu.pl
Hayward
B
B
Borek
D
D
Bonthron
D T
DT
Jaskólski
M
M
D003460:Crystallization; D004926:Escherichia coli; D005631:Fructokinases / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D006801:Humans; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D014961:X-Ray Diffraction
D011994:Recombinant Proteins; D005631:Fructokinases; C021475:ketohexokinase