Purification and characteristics of functional properties of soluble nucleoside triphosphatase (apyrase) from bovine brain.
Soluble NTPase, differing in its properties from known proteins exhibiting NTPase activity, was purified from bovine brain to homogeneity. The enzyme has pH optimum at 7.5 and shows absolute dependence on bivalent cations and broad substrate specificity towards nucleoside-5 -tri- and -diphosphates, characteristics of apyrases. The NTPase follows Michaelis-Menten kinetics in the range of investigated substrate concentrations, the apparent K(m) values for UTP, ITP, GTP, CTP, CDP, and ATP being 86, 25, 41, 150, 500, and 260 microM, respectively. According to gel-filtration and SDS-PAGE data, the molecular mass of the enzyme is 60 kD. The NTPase is localized in the cytosol fraction and expressed in different bovine organs and tissues. Total NTPase activity of extracts of bovine organs and tissues decreases in the following order: liver > heart > skeletal muscle > lung > brain > spleen > kidney ~ small intestine. The enzyme activity can be regulated by acetyl-CoA, alpha-ketoglutarate, and fructose-1,6-diphosphate acting as activators in physiological concentrations, whereas propionate exhibits an inhibitory effect.
Sivuk
V F
VF
Laboratory of Biochemical Toxicology and Narcology, Institute of Pharmacology and Biochemistry, National Academy of Sciences of Belarus, Grodno, 230030, Belarus.
Rusina
I M
IM
Makarchikov
A F
AF
D000105:Acetyl Coenzyme A / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001081:Apyrase / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D001921:Brain / Q000201:enzymology*; D002412:Cations / Q000378:metabolism; D002417:Cattle; D003570:Cytidine Triphosphate / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D005635:Fructosediphosphates / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D007293:Inosine Triphosphate / Q000378:metabolism; D007668:Kidney / Q000201:enzymology; D007700:Kinetics; D008099:Liver / Q000201:enzymology; D043583:Nucleoside-Triphosphatase / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D011422:Propionates / Q000378:metabolism; D013379:Substrate Specificity; D014544:Uridine Triphosphate / Q000378:metabolism
D002412:Cations; D005635:Fructosediphosphates; D011422:Propionates; D007293:Inosine Triphosphate; D003570:Cytidine Triphosphate; D000105:Acetyl Coenzyme A; D006160:Guanosine Triphosphate; D000255:Adenosine Triphosphate; D043583:Nucleoside-Triphosphatase; D001081:Apyrase; C029658:propionic acid; C029063:fructose-1,6-diphosphate; D014544:Uridine Triphosphate