An NAD +-dependent glutamate dehydrogenase cloned from the ruminal ciliate protozoan, Entodinium caudatum

• Published in: FEMS microbiology letters Vol. 247; no. 2; pp. 113 - 121
• Main Authors: Newbold, C. James, McEwan, Neil R., Calza, Roger E., Chareyron, Emilie N., Duval, Stéphane M., Eschenlauer, Sylvain C.P., McIntosh, Freda M., Nelson, Nancy, Travis, Anthony J., Wallace, R. John
• Format: Journal Article
• Language: English
• Published: Oxford, UK Elsevier B.V 15-06-2005; Blackwell Publishing Ltd; Blackwell; Oxford University Press
• Subjects: Affinity; Amino Acid Sequence; Ammonia; Ammonia - metabolism; Ammonia uptake; Animals; Antibiotics; Bacteria; Bacteriology; Bacteroides - genetics; Base Sequence; Biological and medical sciences; Cattle; Ciliophora - enzymology; Cloning; Cloning, Molecular; Dehydrogenase; Dehydrogenases; DNA, Protozoan - chemistry; DNA, Protozoan - genetics; E coli; Entodinium caudatum; Enzymes; Escherichia coli - genetics; Escherichia coli - metabolism; Fundamental and applied biological sciences. Psychology; Gene Expression Regulation; Gene transfer; Gene Transfer, Horizontal; Genes, Protozoan; Genetics; Glutamate dehydrogenase; Glutamate Dehydrogenase - genetics; Glutamate Dehydrogenase - isolation & purification; Glutamate Dehydrogenase - metabolism; Glutamic Acid - metabolism; Ketoglutaric acid; Ketoglutaric Acids - metabolism; Metabolism. Enzymes; Microbiology; Molecular Sequence Data; mRNA; NAD; Phylogeny; Protozoa; Recombinant Proteins - metabolism; RNA, Messenger - analysis; RNA, Protozoan - analysis; Rumen; Sequence Homology, Amino Acid; Substrate Specificity; Glutamate dehydrogenase; Protozoa; Rumen; Entodinium caudatum; Ammonia uptake; Enzyme; Phylogenetic tree; Escherichia coli; Ciliated cell; Uptake; Ammonia; Enzymatic activity; Bacteria; Oxidoreductases; Bacteroid; Enterobacteriaceae
• ISSN: 0378-1097; 1574-6968
• DOI: 10.1016/j.femsle.2005.04.034

An NAD
+-dependent glutamate dehydrogenase (GDH; EC 1.4.1.24) was cloned from the ruminal ciliate protozoan,
Entodinium caudatum. The gene had high sequence similarity to GDH genes from the Bacteroides (class) – a class of bacteria which is highly represented in the rumen. When expressed in
Escherichia coli the enzyme had a high affinity for ammonia and α-ketoglutarate (apparent
K
m of 2.33 and 0.71
mM, respectively) and a low affinity for glutamate (apparent
K
m of 98
mM). GDH activity and GDH mRNA concentration were increased by incubating washed
E. caudatum cells with ammonia and antibiotics. These results suggest that the GDH is an anabolic enzyme catalysing the assimilation of ammonia by
E. caudatum in the rumen and that the gene was probably acquired by lateral gene transfer from a ruminal bacterium.