文摘
Changes in the levels of ferredoxin (Fd)-dependent glutamate synthase (EC 1.4.7.1) and its mRNA were investigated in leaves of maize (Zea mays L. cv DEA). In etiolated leaves, detached from nitrogen-starved dark-grown seedlings, Fd-glutamate synthase was present at a low level. The enzyme protein and the activity were induced from 3- to 5-fold during the 35 h after transfer of etiolated leaves to a medium containing either 10 mM KNO3, 10 mM NH4Cl or 10 mM NH4NO3 under the continuous photosynthetic photon flux density of 300 μmol quanta/m2 per s. A slight increase in the activity occurred in the leaves grown in a nitrogen-free medium under continuous light. The increase in the enzyme activity was paralleled by the incorporation of l-[35S]methionine in the medium into the Fd-glutamate synthase polypeptide under the same conditions. The production of the enzyme protein and the uptake of labeled amino acid into the enzyme protein were blocked by adding 71 μM cycloheximide to the medium supplemented with KNO3, NH4Cl or NH4NO3. The addition of the different nitrogenous compounds under continuous darkness did not significantly alter the Fd-glutamate synthase activity and l-[35S]methionine incorporation into the enzyme protein. A partial cDNA of 2479 base pairs long encoding maize Fd-glutamate synthase was cloned and characterized. Using this cDNA as hybridization probe, Fd-glutamate synthase mRNA was observed to be present at a low level in nitrogen-starved etiolated leaves. The mRNA increased about 5-fold when etiolated leaves were incubated under continuous light in a medium containing either 10 mM KNO3, 10 mM NH4Cl or 10 mM NH4NO3. The level of mRNA was also slightly enhanced in the leaves incubated in a nitrogen-free medium under the light. The exogenous nitrogen compounds did not increase the level of the mRNA under continuous darkness. The presence of 71 μM cycloheximide in either of the media did not significantly change the level of the transcript during the initial 6 h. The induction of mRNA in the presence of exogenous nitrogen under the light was consistent with a protein synthesis-independent process in maize leaves.
