title: Properties and molecular evolution of human GLUD2 (neural and testicular tissue-specific) glutamate dehydrogenase.
abstract: Glutamate dehydrogenase (GDH) is an enzyme central to the metabolism of glutamate that also plays a role in cellular energetics. In the human, GDH exists in a housekeeping isoenzyme (hGDH1) encoded by the GLUD1 gene and a neural and testicular tissue-specific isoform (hGDH2) encoded by the GLUD2 gene. There is evolutionary evidence that the GLUD1 was retroposed to the X chromosome in the ape ancestor (>23 million years ago), where it gave rise to GLUD2 through random mutations and directional selection. In the human, the two mature GDH isoproteins are highly homologous, differing in only 16 of their 505 amino acid residues. Functional analyses of highly purified recombinant wild-type hGDH2 revealed that this adaptive evolution dissociated the enzyme from GTP control, permitted regulation almost entirely by ADP and/or L-leucine, and fine-tuned its activity to the relatively low cellular pH that occurs in synaptic astrocytes during excitatory transmission. Study of structure-function relationships, using site-directed mutagenesis of GLUD1 at single sites differing from GLUD2, showed that the Arg443Ser and the Gly456Ala change reproduced some, but not all, of the properties of hGDH2. In addition, we created a double hGDH1 mutant that had both Arg443Ser and Gly456Ala in the same polypeptide chain. Functional analyses revealed that the doubly mutated enzyme did not acquire all the characteristics of the wild-type hGDH2. Hence, additional amino acid changes, acting in concert with Arg443Ser and Gly456Ala, ought to be responsible the unique properties of the brain-specific human isoenzyme.
authors.affiliation: Department of Neurology, University of Crete School of Health Sciences, Section of Medicine, Heraklion, Crete, Greece.
authors.forename: Konstantinos
authors.initials: K
authors.lastname: Kanavouras
authors.forename: Vasileios
authors.initials: V
authors.lastname: Mastorodemos
authors.forename: Nikolas
authors.initials: N
authors.lastname: Borompokas
authors.forename: Cleanthe
authors.initials: C
authors.lastname: Spanaki
authors.forename: Andreas
authors.initials: A
authors.lastname: Plaitakis
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D004591:Electrophoresis, Polyacrylamide Gel; D019143:Evolution, Molecular; D005969:Glutamate Dehydrogenase / Q000235:genetics*; D006801:Humans; D007527:Isoenzymes / Q000235:genetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D013329:Structure-Activity Relationship
chemical_list: D007527:Isoenzymes; D005969:Glutamate Dehydrogenase; C521120:GLUD2 protein, human