Open AccessNADP + ‐dependent glutamate dehydrogenase from the facultative methylotroph Hyphomicrobium X
Author(s) -
Duchars M.G.,
Attwood Margaret M.
Publication year - 1987
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.1987.tb02529.x
Subject(s) - glutamate dehydrogenase , methylotroph , chromatography , glutamate synthase , chemistry , ammonium , enzyme , polyacrylamide gel electrophoresis , gel electrophoresis , biochemistry , sodium , glutamate receptor , receptor , organic chemistry
NADP + ‐dependent glutamate dehydrogenase (GDH; EC 1.4.1.4) was purified using acetone precipitation, heat, DEAE‐cellulose and dye‐ligand Ramazol Red column chromatography. The M r of the native enzyme was estimated to be 380 000 (± 10 000) by polyacrylamide gel electrophoresis. The same technique in the presence of sodium dodecyl sulphate (SDS) gave one subunit band with an M r of 63 400 (±4000). Thus the enzyme has a hexameric structure. The enzyme has a pH optimum of 8.5 and has K m apparent values of 1.6 mM, 0.015 mM and 10.2 mM for α‐ketoglutarate, N NADPH and L ‐glutamate, respectively. Michaelis‐Menten kinetics were not observed when the ammonium concentration was increased. A progressive increase in the ammonium concentration resulted in a progressively increasing K m value. The enzyme was highly specific for all substrates and markedly insensitive to inhibitors.