PMID:22068497
| Citation |
Wang, B, Wang, P, Zheng, E, Chen, X, Zhao, H, Song, P, Su, R, Li, X and Zhu, G (2011) Biochemical properties and physiological roles of NADP-dependent malic enzyme in Escherichia coli. J. Microbiol. 49:797-802 |
|---|---|
| Abstract |
Malic enzymes catalyze the reversible oxidative decarboxylation of L-malate using NAD(P)(+) as a cofactor. NADP-dependent malic enzyme (MaeB) from Escherichia coli MG1655 was expressed and purified as a fusion protein. The molecular weight of MaeB was about 83 kDa, as determined by SDS-PAGE. The recombinant MaeB showed a maximum activity at pH 7.8 and 46°C. MaeB activity was dependent on the presence of Mn(2+) but was strongly inhibited by Zn(2+). In order to understand the physiological roles, recombinant E. coli strains (icd (NADP)/ΔmaeB and icd (NAD)/ΔmaeB) containing NADP-dependent isocitrate dehydrogenase (IDH), or engineered NAD-dependent IDH with the deletion of the maeB gene, were constructed using homologous recombination. During growth on acetate, icd (NAD)/ΔmaeB grew poorly, having a growth rate only 60% that of the wild-type strain (icd (NADP)). Furthermore, icd (NADP)/ΔmaeB exhibited a 2-fold greater adaptability to acetate than icd (NAD)/ΔmaeB, which may be explained by more NADPH production for biosynthesis in icd (NADP)/ΔmaeB due to its NADP-dependent IDH. These results indicated that MaeB was important for NADPH production for bacterial growth on acetate. We also observed that MaeB activity was significantly enhanced (7.83-fold) in icd (NAD), which was about 3-fold higher than that in icd (NADP), when switching from glucose to acetate. The marked increase of MaeB activity was probably induced by the shortage of NADPH in icd (NAD). Evidently, MaeB contributed to the NADPH generation needed for bacterial growth on two carbon compounds. |
| Links |
PubMed Online version:10.1007/s12275-011-0487-5 |
| Keywords |
Acetates/metabolism; Coenzymes/metabolism; Electrophoresis, Polyacrylamide Gel; Enzyme Activators/metabolism; Enzyme Inhibitors/metabolism; Enzyme Stability; Escherichia coli/enzymology; Escherichia coli/genetics; Escherichia coli/growth & development; Escherichia coli/physiology; Gene Expression; Hydrogen-Ion Concentration; Malate Dehydrogenase/genetics; Malate Dehydrogenase/metabolism; Manganese/metabolism; Metabolic Engineering; Metabolic Networks and Pathways/genetics; Molecular Weight; NADP/metabolism; Recombinant Fusion Proteins/chemistry; Recombinant Fusion Proteins/genetics; Recombinant Fusion Proteins/isolation & purification; Recombinant Fusion Proteins/metabolism; Temperature; Zinc/metabolism |
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