title: An NAD(+)-dependent glutamate dehydrogenase cloned from the ruminal ciliate protozoan, Entodinium caudatum.
abstract: 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 alpha-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.
authors.affiliation: Rowett Research Institute, Greenburn Road, Aberdeen AB21 9SB, Scotland, United Kingdom.
authors.forename: C James
authors.initials: CJ
authors.lastname: Newbold
authors.forename: Neil R
authors.initials: NR
authors.lastname: McEwan
authors.forename: Roger E
authors.initials: RE
authors.lastname: Calza
authors.forename: Emilie N
authors.initials: EN
authors.lastname: Chareyron
authors.forename: Stéphane M
authors.initials: SM
authors.lastname: Duval
authors.forename: Sylvain C P
authors.initials: SC
authors.lastname: Eschenlauer
authors.forename: Freda M
authors.initials: FM
authors.lastname: McIntosh
authors.forename: Nancy
authors.initials: N
authors.lastname: Nelson
authors.forename: Anthony J
authors.initials: AJ
authors.lastname: Travis
authors.forename: R John
authors.initials: RJ
authors.lastname: Wallace
mesh_terms: D000595:Amino Acid Sequence; D000641:Ammonia / Q000378:metabolism; D000818:Animals; D001439:Bacteroides / Q000235:genetics; D001483:Base Sequence; D016798:Ciliophora / Q000201:enzymology*; D003001:Cloning, Molecular; D016054:DNA, Protozoan / Q000737:chemistry / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation; D022761:Gene Transfer, Horizontal; D017125:Genes, Protozoan; D005969:Glutamate Dehydrogenase / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D018698:Glutamic Acid / Q000378:metabolism; D007656:Ketoglutaric Acids / Q000378:metabolism; D008969:Molecular Sequence Data; D010802:Phylogeny; D012333:RNA, Messenger / Q000032:analysis; D016053:RNA, Protozoan / Q000032:analysis; D011994:Recombinant Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
chemical_list: D016054:DNA, Protozoan; D007656:Ketoglutaric Acids; D012333:RNA, Messenger; D016053:RNA, Protozoan; D011994:Recombinant Proteins; D018698:Glutamic Acid; D000641:Ammonia; D005969:Glutamate Dehydrogenase