title: Kinetic and chemical mechanisms for the effects of univalent cations on the spectral properties of aromatic amine dehydrogenase.
abstract: Univalent cations and pH influence the UV-visible absorption spectrum of the tryptophan tryptophylquinone (TTQ) enzyme, aromatic amine dehydrogenase (AADH). Little spectral perturbation was observed when pH was varied in the absence of univalent cations. The addition of alkali metal univalent cations (K+, Na+, Li+, Rb+ or Cs+) to oxidized AADH caused significant changes in its absorption spectrum. The apparent Kd for each cation, determined from titrations of the spectral perturbation, decreased with increasing pH. Transient kinetic studies involving rapid mixing of AADH with cations and pH jump revealed that the rate of the cation-induced spectral changes initially decreased with increasing cation concentration to a minimum value, then increased with increasing cation concentration. A kinetic model was developed to fit these data, determine the true pH-independent Kd values for K+ and Na+, and explain the pH-dependence of the apparent Kd. A chemical reaction mechanism, based on the kinetic data, is presented in which the metallic univalent cation facilitates the chemical modification of the TTQ prosthetic group to form an hydroxide adduct which gives rise to the spectral change. Addition of NH4(+)/NH3 to AADH caused changes in the absorption spectrum which were very different form those caused by addition of the metallic univalent cations. The kinetics of the reaction induced by addition of NH4+/NH3 were also different, being simple saturation kinetics. Another reaction mechanism is proposed for the NH4+/NH3-induced spectral change that involves nucleophilic addition of the unprotonated NH3 to TTQ. The general relevance of these data and models to the physiological reactions of TTQ-dependent enzymes and to the roles of univalent cations in modulating enzyme activity are discussed.
authors.affiliation: Department of Biochemistry, University of Mississippi Medical Center, Jackson, MS 39216-4505, USA.
authors.forename: Z
authors.initials: Z
authors.lastname: Zhu
authors.forename: V L
authors.initials: VL
authors.lastname: Davidson
mesh_terms: D000421:Alcaligenes / Q000201:enzymology*; D000641:Ammonia / Q000378:metabolism; D002414:Cations, Monovalent / Q000378:metabolism / Q000494:pharmacology*; D006863:Hydrogen-Ion Concentration; D045563:Indolequinones; D007700:Kinetics; D008672:Metals, Alkali / Q000378:metabolism / Q000494:pharmacology*; D015394:Molecular Structure; D000587:Oxidoreductases Acting on CH-NH Group Donors / Q000737:chemistry* / Q000378:metabolism; D011188:Potassium / Q000378:metabolism / Q000494:pharmacology; D011485:Protein Binding; D011809:Quinones / Q000378:metabolism; D012964:Sodium / Q000378:metabolism / Q000494:pharmacology; D013053:Spectrophotometry; D014364:Tryptophan / Q000031:analogs & derivatives / Q000378:metabolism
chemical_list: D002414:Cations, Monovalent; D045563:Indolequinones; D008672:Metals, Alkali; D011809:Quinones; C070569:tryptophan tryptophylquinone; D000641:Ammonia; D014364:Tryptophan; D012964:Sodium; D000587:Oxidoreductases Acting on CH-NH Group Donors; C037193:aromatic amine dehydrogenase; D011188:Potassium