Collect. Czech. Chem. Commun.
2007, 72, 1420-1434
https://doi.org/10.1135/cccc20071420
The Enzyme Kinetics of the NADP-Malic Enzyme from Tobacco Leaves
Helena Ryšlaváa,*, Veronika Doubnerováa, Karel Mullera, Petra Baťkováb,c, Renáta Schnablováb,c, Jiří Liberdaa, Helena Synkováb and Noemi Čeřovskáb
a Department of Biochemistry, Faculty of Science, Charles University, Hlavova 2030, CZ-128 40 Prague 2, Czech Republic
b Institute of Experimental Botany, Academy of Sciences of the Czech Republic, Na Karlovce 1a, CZ-160 00 Prague 6, Czech Republic
c Department of Plant Anatomy and Physiology, Faculty of Science, Charles University, Viničná 5, CZ-128 44 Prague 2, Czech Republic
Abstract
Malic enzyme (L-malate: NADP+ oxidoreductase (oxaloacetate-decarboxylating), EC 1.1.1.40, NADP-ME), which was found in chloroplasts, was isolated from tobacco leaves (Nicotiana tabacum L.) almost homogenous. The specific enzyme activity was 0.95 μmol min-1 mg-1. The enzyme pH optimum was found between pH 7.1 and 7.4. The affinity of NADP-ME to substrates (L-malate and NADP+) was evaluated in the presence of divalent metal ions (Mg2+, Mn2+, Co2+, Ni2+). The value of the apparent Michaelis constant of NADP-ME for L-malate was dependent on the ion cofactor, while no such relationship was found for NADP+. The dependence of the reaction rate on concentration of Mg2+ indicates the presence of more than one binding site for these ions in NADP-ME. Likewise, the sigmoidal dependence of the reaction rate on Mn2+ concentration and the value of Hill coefficient 7.5 indicate the positive cooperativity of the reaction kinetics in the presence of the ions. The effect of Co2+ and Ni2+ ions was analogous to that of Mn2+ ions; however, the cooperativity was lower (the values of Hill coefficients were 3.0 and 1.3 for Co2+ and Ni2+, respectively). Regulation of NADP-ME from tobacco leaves by divalent metal ions is discussed.
Keywords: Oxidoreductases; Enzyme kinetics; NADP-malic enzyme; Divalent metal ions; Nicotiana tabacum L.
References: 31 live references.
