title: Molecular and biochemical characterisation of a Teladorsagia circumcincta glutamate dehydrogenase.
abstract: A full length cDNA encoding glutamate dehydrogenase was cloned from Teladorsagia circumcincta (TcGDH). The TcGDH cDNA (1614 bp) encoded a 538 amino acid protein. The predicted amino acid sequence showed 96% and 93% similarity with Haemonchus contortus and Caenorhabditis elegans GDH, respectively. A soluble N-terminal 6xHis-tagged GDH protein was expressed in the recombinant Escherichia coli strain BL21 (DE3) pGroESL, purified and characterised. The recombinant TcGDH had similar kinetic properties to those of the enzyme in homogenates of T. circumcincta, including greater activity in the aminating than deaminating reaction. Addition of 1mM ADP and ATP increased activity about 3-fold in the deaminating reaction, but had no effect in the reverse direction. TcGDH was a dual co-factor enzyme that operated both with NAD(+) and NADP(+), GDH activity was greater in the deaminating reaction with NADP(+) as co-factor and more with NADH in the aminating reaction.
authors.affiliation: Institute of Veterinary, Animal and Biomedical Sciences, Massey University, Private Bag 11-222, Palmerston North, New Zealand.
authors.forename: S
authors.initials: S
authors.lastname: Umair
authors.forename: J S
authors.initials: JS
authors.lastname: Knight
authors.forename: M L
authors.initials: ML
authors.lastname: Patchett
authors.forename: R J
authors.initials: RJ
authors.lastname: Bland
authors.forename: H V
authors.initials: HV
authors.lastname: Simpson
mesh_terms: D000586:Amination; D000595:Amino Acid Sequence; D000641:Ammonia / Q000378:metabolism; D000818:Animals; D018076:DNA, Complementary / Q000737:chemistry / Q000302:isolation & purification; D018509:DNA, Helminth / Q000737:chemistry* / Q000302:isolation & purification; D003641:Deamination; D015971:Gene Expression Regulation, Enzymologic; D005969:Glutamate Dehydrogenase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D018698:Glutamic Acid / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D009243:NAD / Q000378:metabolism; D009249:NADP / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D012756:Sheep; D014251:Trichostrongyloidea / Q000201:enzymology* / Q000235:genetics
chemical_list: D018076:DNA, Complementary; D018509:DNA, Helminth; D011994:Recombinant Proteins; D009243:NAD; D018698:Glutamic Acid; D009249:NADP; D000641:Ammonia; D005969:Glutamate Dehydrogenase