Isolation and properties of glyceraldehyde-3-phosphate dehydrogenase from a sturgeon from the Caspian Sea and its interaction with spin-labeled NAD+ derivatives.
Glyceraldehyde-3-phosphate dehydrogenase (D-glyceraldehyde-3-phosphate: NAD+ oxidoreductase (phosphorylating), EC 1.2.1.12) was isolated from a sturgeon, Huso huso, from the Caspian Sea. It is closely related to the enzyme from a Pacific sturgeon, Acipenser transmontanus, with respect to amino acid composition, steady-state kinetics and coenzyme binding. The latter, as studied by means of a spin-labeled derivative of NAD+, is negatively cooperative exhibiting a Hill coefficient of 0.84 at 12 degrees C. Two derivatives of NAD+ spin-labeled at N6 or C8 of the adenine ring were found to be active coenzymes with maximum velocities reaching 35 or 45% of the value for NAD+ itself. When more than two equivalents of either spin-labeled NAD+ are bound to the enzyme spin-spin interactions are observed in the ESR spectra. Distances between the nitroxide radicals (8--9 A) calculated from the observed splittings are in excellent agreement with data predicted from the crystal structure of the lobster enzyme when the coenzyme is bound in an anti-conformation of the adenine moiety about the glycosidic bond to all four subunits.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002852:Chromatography, Ion Exchange; D004578:Electron Spin Resonance Spectroscopy; D005399:Fishes; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000302:isolation & purification* / Q000378:metabolism; D009243:NAD / Q000378:metabolism*; D013113:Spin Labels
D000596:Amino Acids; D013113:Spin Labels; D009243:NAD; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases