Characterization of alpha-phosphoglucomutase isozymes from Toxoplasma gondii.
The Toxoplasma gondii genome project has revealed two putative isoforms (TgPGM-I and TgPGM-II) of alpha-phosphoglucomutase (EC 5.4.2.2). We obtained recombinant proteins of these isoforms from the Beverley strain of T. gondii and characterized their properties, particularly the kinetic properties of these isoforms. The specific activities of TgPGM-I and TgPGM-II for alpha-D-glucose 1-phosphate were 338+/-9 and 84+/-6micromol/min/mg protein, respectively, at 37 degrees C under optimal conditions. The Kcat and Km values of TgPGM-I were 398+/-11/s and 0.19+/-0.03mM and those for TgPGM-II were 93+/-7/s and 3.53+/-0.91mM, respectively, for alpha-d-glucose 1-phosphate. Magnesium ions were the most effective divalent cations for both the enzyme activities. The maximum activities of both the enzymes were obtained in the presence of more than 0.2mM alpha-D-glucose 1,6-bisphosphate. Although both enzymes were attached to the alpha-phosphohexomutase superfamily, amino acid sequence homology between TgPGM-I and TgPGM-II showed very low overall identity (25%). No alpha-phosphomannomutase (EC 5.4.2.8) activity was detected for either enzyme. The data indicated that TgPGM-I, but not TgPGM-II, may play an important role in alpha-D-glucose 6-phosphate production.
Department of Tropical Medicine and Parasitology, School of Medicine, Keio University, Shinjuku-ku, Tokyo, Japan.
Mihoko
M
Imada
Shuichi
S
Kawashima
Minoru
M
Kanehisa
Tsutomu
T
Takeuchi
Takashi
T
Asai
D000595:Amino Acid Sequence; D000818:Animals; D019298:Glucose-6-Phosphate / Q000378:metabolism; D006019:Glycolysis; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D010733:Phosphoglucomutase / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D010802:Phylogeny; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D014122:Toxoplasma / Q000201:enzymology* / Q000235:genetics
D007527:Isoenzymes; D011994:Recombinant Proteins; D019298:Glucose-6-Phosphate; D010733:Phosphoglucomutase