title: Room temperature phosphorescence of Trp-314 as a monitor of subunit communications in alcohol dehydrogenase from horse liver.
abstract: The phosphorescence properties of liver alcohol dehydrogenase from horse were characterized at limiting concentrations of coenzyme and coenzyme analogues. The emission decay kinetics of Trp-314 in strong, slowly exchanging, ternary complexes with NADH/isobutyramide, NAD/pyrazole, and NADH/dimethyl sulfoxide displays a markedly nonexponential character. The analysis of decay components over the saturation curve reveals that the phosphorescence from singly bound protein molecules has a lifetime from 1 to 1.3 s, which is 2-3 times larger than observed with fully bound and unliganded enzyme. The remarkably tighter configuration reported by the triplet probe for the coenzyme-binding domain in half-saturated macromolecules is not exclusive of strongly inhibited ternary complexes. Measurements on binary complexes with NADH, ADPR, and the inactive coenzyme analogue 1,4,5,6-tetrahydronicotinamide adenine dinucleotide confirm that binding of the ligand to one subunit has qualitatively the same influence on protein structure. If the lifetime of Trp-314 provides clear evidence for an appreciable change in conformation at half-binding that is apparently triggered by the ADPR fragment of the coenzyme, such communication between subunits does not lead to allosteric phenomena in coenzyme binding.
authors.affiliation: CNR, Istituto di Biofisica, Pisa, Italy.
authors.forename: G B
authors.initials: GB
authors.lastname: Strambini
authors.forename: M
authors.initials: M
authors.lastname: Gonnelli
authors.forename: W C
authors.initials: WC
authors.lastname: Galley
mesh_terms: D000426:Alcohol Dehydrogenase / Q000378:metabolism*; D000577:Amides / Q000378:metabolism; D000818:Animals; D003067:Coenzymes / Q000378:metabolism; D006736:Horses; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D008163:Luminescent Measurements; D015394:Molecular Structure; D009243:NAD / Q000378:metabolism; D010084:Oxidation-Reduction; D011487:Protein Conformation; D013696:Temperature; D014364:Tryptophan / Q000378:metabolism
chemical_list: D000577:Amides; D003067:Coenzymes; D009243:NAD; C030197:isobutyramide; D014364:Tryptophan; D000426:Alcohol Dehydrogenase