Open AccessGlutamate dehydrogenase (NADP‐dependent) mRNA in relation to enzyme synthesis in Euglena gracilis
Author(s) -
PARKER Jane E.,
JAVED Qamar,
MERRETT Michael J.
Publication year - 1985
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1985.tb09339.x
Subject(s) - glutamate dehydrogenase , biochemistry , glutamate receptor , biology , dehydrogenase , glutamate synthase , microbiology and biotechnology , reticulocyte , messenger rna , enzyme , receptor , gene
Cells of Euglena gracilis Klebs strain z Pringsheim had high NADP‐dependent glutamate dehydrogenase activity when grown on glutamate as nitrogen source but activity was completely repressed in cells grown on ammonium (NH + 4 ). A 120‐fold purification of NADPH‐glutamate dehydrogenase (subunit M r = 45000) was achieved from glutamate‐grown cells by affinity chromatography on blue Sepharose CL‐6B. Antisera raised against the homogeneously pure protein were used to demonstrate that increase in NADPH‐glutamate dehydrogenase activity on transfer from NH + 4 ‐grown cells were labelled with L‐[ 35 S]methionine and anti‐(NADPH‐glutamate dehydrogenase) used to immunoprecipitate the dehydrogenase from cell extracts. NADPH‐glutamate dehydrogenase protein was detected in glutamate‐grown but not NH + 4 ‐grown cells. Anti‐(NADPH‐glutamate dehydrogenase) was used to detect NADPH‐glutamate dehydrogenase resulting from the translation of total polyadenylated RNA from Euglena in a cell‐free rabbit reticulocyte lysate system. NADPH‐glutamate dehydrogenase mRNA was present in glutamate NH + 4 ‐grown cells, there being no apparent difference in mRNA abundance between cells showing a tenfold difference in NADPH‐glutamate dehydrogenase specific activity. These results indicate that the synthesis of this dehydrogenase is regulated primarily at the post‐transcriptional level.