Restraint-based comparative modeling was used for calculation and visualization of the H4-H5-loop of Na+/K+-ATPase from mouse brain (Mus musculus, adult male brain, α2-isoform) between the amino acid residues Cys336 and Arg758 in the E1 conformation The structure consists of two well separated parts. The N-domain is formed by a seven-stranded antiparallel β-sheet with two additional β-strands and five α-helices sandwiching it, the P-domain is composed of a typical Rossman fold. The ATP-binding site was found on the N-domain to be identical in both α2- and α1-isoforms. The phosphorylation Asp369 residue was found in the central part of the P-domain, located at the C-terminal end of the central β-sheet. The distance between the α-carbon of Phe475 at the ATP-binding site and the α-carbon of Asp369 at the phosphorylation site is 3.22 nm. A hydrogen bond between the oxygen atom of Asp369 and the nitrogen atom of Lys690 was clearly detected and assumed to play a key role in maintaining the proper structure of the physphorylaton site in E1 conformation., G. Tejral, L. Koláčná, A. Kotyk, E. Amler., and Obsahuje bibliografii
Microsomes were prepared from placentas of normotensive women and of patients suffering from pregnancy- induced hypertension (PIH). Activity of Na,K-ATPase (estimated as ATP hydrolysis) from the hypertensive tissue was lower than from tissue of normotensive women, even if the number of Na,K-ATPase molecules (monitored by anthroyl ouabain binding) was actually greater in the hypertensive tissue. The affinity of Na,K-ATPase for anthroyl ouabain was about four times higher in plasma membranes of hypertensives, indicating some structural change in the Na,K-ATPase or in its vicinity. Assuming the presence of an endogenous digitalis-like factor, the results suggest a simple way of explaining not only the lower Ma,K-ATPase activity in the placental membranes of hypertensives but also the different extent of enzyme inhibition in different tissues of PIH patients.
Ornithine carbamoyltransferase has been purified from the liver of the loggerhead turtle Caretta caretta by a single-step procedure using chromatography on an affinity column to which the transition-state analogue, d-N-(phosphonoacetyl)- L-ornithine (d-PALO), was covalently bound. The procedure employed yielded an enzyme which was purified 373-fold and was judged to be homogeneous by nondenaturing and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The enzyme showed a specific activity of 224. The molar mass of the C. caretta enzyme was approximately 112 kDa, the single band obtained by SDS-PAGE indicated a subunit molar mass of 39.5 kDa; hence, the enzyme is a trimer of identical subunits. It catalyzes an ordered sequential mechanism in which carbamoyl phosphate binds first, followed by L-ornithine. The Michaelis constants were 0.858 mM for L-ornithine and 0.22 mM for carbamoyl phosphate, the dissociation constant of the enzyme-carbamoyl phosphate complex was 0.50 mM., E. Bellocco, C. Di Salvo, G. Lagan, U. Leuzzi, E. Tellone, A. Kotyk, A. Galtieri., and Obsahuje bibliografii