title: Identification of amino acid residues responsible for different GTP preferences of human glutamate dehydrogenase isozymes.
abstract: Human glutamate dehydrogenase isozymes (hGDH1 and hGDH2) differ markedly in their inhibition by GTP. These regulatory preferences must arise from amino acid residues that are not common between hGDH isozymes. We have constructed chimeric enzymes by reciprocally switching the corresponding amino acid segments 390-465 in hGDH isozymes that are located within or near the C-terminal 48-residue antenna helix, which is thought to be part of the regulatory domain of mammalian GDHs. These resulted in triple mutations in amino acid sequences at 415, 443, and 456 sites that are not common between hGDH1 and hGDH2. The chimeric enzymes did not change their enzyme efficiency (k(cat)/K(m)) and expression level. Functional analyses, however, revealed that the chimeric mutants almost completely acquired the different GTP regulatory preference between hGDH isozymes. These results suggest that the 415, 443, and 456 residues acting in concert are responsible for the GTP inhibitory properties of hGDH isozymes.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Ulsan College of Medicine, 388-1 Poongnap-dong, Songpa-gu, Seoul 138-736, Republic of Korea.
authors.forename: Myung-Min
authors.initials: MM
authors.lastname: Choi
authors.forename: Eun Young
authors.initials: EY
authors.lastname: Hwang
authors.forename: Eun-A
authors.initials: EA
authors.lastname: Kim
authors.forename: Jae-Wan
authors.initials: JW
authors.lastname: Huh
authors.forename: Sung-Woo
authors.initials: SW
authors.lastname: Cho
mesh_terms: D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry*; D001665:Binding Sites; D003198:Computer Simulation; D004789:Enzyme Activation; D005969:Glutamate Dehydrogenase / Q000737:chemistry*; D006160:Guanosine Triphosphate / Q000737:chemistry*; D006801:Humans; D007527:Isoenzymes / Q000737:chemistry; D007700:Kinetics; D008956:Models, Chemical; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding
chemical_list: D000596:Amino Acids; D007527:Isoenzymes; D006160:Guanosine Triphosphate; D005969:Glutamate Dehydrogenase