title: Novel allosteric properties produced by residue substitutions in the subunit interface of yeast NAD+-specific isocitrate dehydrogenase.
abstract: Yeast NAD+-specific isocitrate dehydrogenase (IDH) is an octamer of four IDH1 and four IDH2 subunits, and the basic structural unit of the enzyme is an IDH1/IDH2 heterodimer. To investigate one aspect of the interaction between IDH1 and IDH2, residues in a hydrophobic region at the heterodimer interface (Val-216, Ser-220, and Val-224 in IDH1; Ile-221, Val-225, and Val-229 in IDH2) were replaced by alanine residues in each and in both subunits. Gel filtration and sedimentation velocity analyses demonstrated that the residue substitutions do not disrupt the octameric structure of IDH. However, these substitutions produce novel kinetic properties including, with respect to cofactor, positive allosteric regulation by AMP and cooperativity in the absence of AMP. These allosteric properties are also apparent in NAD+-binding experiments. Despite substantial measurable activity for the mutant enzyme containing residue substitutions in both subunits, expression of this enzyme produces growth phenotypes indicative of IDH dysfunction in vivo.
authors.affiliation: Department of Biochemistry, University of Texas Health Science Center, San Antonio, TX 78229-3900, USA.
authors.forename: Gang
authors.initials: G
authors.lastname: Hu
authors.forename: Lee
authors.initials: L
authors.lastname: McAlister-Henn
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000596:Amino Acids / Q000737:chemistry*; D003198:Computer Simulation; D007521:Isocitrate Dehydrogenase / Q000737:chemistry* / Q000648:ultrastructure*; D007536:Isomerism; D008956:Models, Chemical; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D021122:Protein Subunits; D013329:Structure-Activity Relationship; D015003:Yeasts / Q000201:enzymology*
chemical_list: D000596:Amino Acids; D021122:Protein Subunits; D007521:Isocitrate Dehydrogenase