Kinetic properties of N6-(2-carboxyethyl)-NAD(H) and poly(ethylene glycol)-bound NAD(H) for alcohol, lactate, malate and glyceraldehyde-3-phosphate dehydrogenase from different organisms

The steady-state kinetics of alcohol dehydrogenases (alcohol:NAD + oxidoreductase, EC 1.1.1.1 and alcohol:NADP + oxidoreductase, EC 1.1.1.2), lactate dehydrogenases ( l -lactate:NAD + oxidoreductase, EC 1.1.1.27 and d -lactate:NAD + oxidoreductase, EC 1.1.1.28), malate dehydrogenase ( l -malate:NAD + oxidoreductase, EC 1.1.1.37), and glyceraldehyde-3-phosphate dehydrogenases [ d -glyceraldehyde-3-phosphate:NAD + oxidoreductase (phosphorylating), EC 1.2.1.12] from different sources (prokaryote and eukaryote, mesophilic and thermophilic organisms) have been studied using NAD(H), N 6 -(2-carboxyethyl)-NAD(H), and poly(ethylene glycol)-bound NAD(H) as coenzymes. The kinetic constants for NAD(H) were changed by carboxyethylation of the 6-amino group of the adenine ring and by conversion to macromolecular form. Enzymes from thermophilic bacteria showed especially high activities for the derivatives. The relative values of the maximum velocity ( NAD = 1) of Thermus thermophilus malate dehydrogenase for N 6 -(2-carboxyethyl)-NAD and poly(ethylene glycol)-bound NAD were 5.7 and 1.9, respectively, and that of Bacillus stearothermophilus glyceraldehyde-3-phosphate dehydrogenase for poly(ethylene glycol)-bound NAD was 1.9.