1887

Abstract

Bf1 possesses two enzymes having glutamate dehydrogenase (GDH) activity. One is dual cofactor NAD(P)H-dependent, while the other has NADH-specific activity. The gene encoding the NADH-GDH () was cloned by complementation of the glutamate auxotrophic mutant MX3004 and the recombinant protein was characterized with respect to the GDH activities present in the parental organism grown under different nitrogen conditions. The NAD(P)H-dependent GDH of was confirmed to be most active under high ammonia conditions, but the NADH-specific GDH levels were increased by high peptide concentrations in the growth medium and not regulated by the levels of ammonia. Northern blotting analysis showed that regulation was at the transcription level, with a single transcript of ∼ 1.6 kb being produced. GDH activity was demonstrated by zymography of the parental and recombinant enzymes. The recombinant GDH was NADH-specific and co-migrated with the equivalent enzyme band from cell extracts. The structural gene comprises 1335 bp and encodes a protein of 445 aa (49 kDa). Comparisons of the derived protein sequence with that of GDH from other bacteria indicated that significant sequence homology and conservation of functional domains exists with enzymes of Family I-type hexameric GDH proteins.

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1998-06-01
2021-10-25
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