Purification and characterization of thiamine triphosphatase from bovine brain.
Soluble thiamine triphosphatase (EC 3.6.1.28) of bovine brain has been purified 68,000-fold to an electrophoretically homogeneous state with an overall recovery of 5.5% by hydrophobic chromatography on Toyopearl HW-60, Sephadex G-75 gel filtration, DEAE-Toyopearl 650M chromatography and Blue Sepharose CL-4B chromatography. The enzyme has an absolute specificity among thiamine and nucleoside phosphate esters for thiamine triphosphate and shows no nonspecific phosphatase activities. Thiamine triphosphatase is composed of a single polypeptide chain with molecular mass of 33,900 kDa as estimated by Sephadex G-100 gel filtration and SDS-polyacrylamide gel electrophoresis. The enzyme has a pH optimum of 8.7 and is dependent on divalent metal ions. Mg2+ has been found to be the most effective among cations tested. A study of the reaction kinetics over a wide range of thiamine triphosphate concentrations has revealed a biphasic saturation curve being described by higher-degree rational polynomials.
Makarchikov
A F
AF
Laboratory of Applied Enzymology and Biotechnology, Academy of Sciences of Belarus, Grodno.
Chernikevich
I P
IP
D000818:Animals; D001921:Brain / Q000201:enzymology*; D002417:Cattle; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008670:Metals; D008970:Molecular Weight; D013379:Substrate Specificity; D013837:Thiamin-Triphosphatase / Q000302:isolation & purification* / Q000378:metabolism
D008670:Metals; D013837:Thiamin-Triphosphatase