title: Kinetic studies on Na+/K+-ATPase and inhibition of Na+/K+-ATPase by ATP.
abstract: Na+/K+-ATPase (EC 3.6.1.3) is an important membrane-bound enzyme. In this paper, kinetic studies on Na+/K+-ATPase were carried out under mimetic physiological conditions. By using microcalorimeter, a thermokinetic method was employed for the first time. Compared with other methods, it provided accurate measurements of not only thermodynamic data (deltarHm) but also the kinetic data (Km and Vmax). At 310.15K and pH 7.4, the molar reaction enthalpy (deltarHm) was measured as -40.514 +/- 0.9kJmol(-1). The Michaelis constant (Km) was determined to be 0.479 +/- 0.020 mM and consistent with literature data. The reliability of the thermokinetic method was further confirmed by colorimetric studies. Furthermore, a simple and reliable kinetic procedure was presented for ascertaining the true substrate for Na+/K+-ATPase and determining the effect of free ATP. Results showed that the MgATP complex was the real substrate with a Km value of about 0.5mM and free ATP was a competitive inhibitor with a Ki value of 0.253 mM.
authors.affiliation: College of Chemistry and Molecular Sciences, Wuhan University, Wuhan 430072, Hubei, People's Republic of China.
authors.forename: Li
authors.initials: L
authors.lastname: Xia
authors.forename: Liu
authors.initials: L
authors.lastname: Yuwen
authors.forename: Li
authors.initials: L
authors.lastname: Jie
authors.forename: Li
authors.initials: L
authors.lastname: Huilin
authors.forename: Yang
authors.initials: Y
authors.lastname: Xi
authors.forename: Wang
authors.initials: W
authors.lastname: Cunxin
authors.forename: Wang
authors.initials: W
authors.lastname: Zhiyong
mesh_terms: D000255:Adenosine Triphosphate / Q000494:pharmacology*; D000818:Animals; D002151:Calorimetry / Q000379:methods; D004789:Enzyme Activation / Q000187:drug effects; D007700:Kinetics; D015636:Magnesium Chloride / Q000737:chemistry / Q000494:pharmacology; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism*; D013552:Swine; D013816:Thermodynamics
chemical_list: D015636:Magnesium Chloride; D000255:Adenosine Triphosphate; D000254:Sodium-Potassium-Exchanging ATPase