[Purification and properties of aromatic L-amino acid decarboxylase (4.1.1.28) of rat brain].
L-aromatic aminoacid decarboxylase has been purified more than thousand times from homogenates of rat brain, in several steps : centrifugation, DEAE-cellulose, CM cellulose, hydroxylapatite, DEAE sephadex. Its properties have been studied, most of them on an intermediate fraction of the purification, because of the instability of the purified enzyme in spite of the addition of different stabilizing agents : the enzyme decarboxylates 5-hydroxytryptophan (5 HTP) and DOPA in a ratio constant throughout the purification but does not decarboxylate tryptophan, tyrosine, histidine at a measurable rate. Optimum pH, Km, Vm, have been measured with 5 HTP and DOPA as substrates. The enzyme has a molecular weight of 115.000, an apparent isoelectric point of 6,4-6,5. It is inhibited by serotonin, dopamine, some cations : Cu++, Fe++, Ni++ by N-ethylmaleimide, sodium dodecylsulfate. Some pyridoxal-5 phosphate (PLP) remains strongly bound to the enzyme. For relatively weak concentrations of substrate, the enzyme is inhibited by an excess of PLP ; for weak concentrations of PLP, the enzyme in inhibited by an excess of substrate, particularly of DOPA. We also observe a spontaneous decarboxylation of the substrates that reaches a plateau and is enhanced by high concentrations of PLP, by serotonin, dopamine, Cu++ and reduced by mercaptoethanol and the presence of crude or boiled homogenates. Several possible explanations of the spontaneous decarboxylation and of the enzymic inhibitions by an excess of PLP and by the substrates are given.
D006916:5-Hydroxytryptophan / Q000378:metabolism; D000818:Animals; D001921:Brain / Q000201:enzymology*; D003300:Copper / Q000494:pharmacology; D004295:Dihydroxyphenylalanine / Q000378:metabolism; D004296:Dopa Decarboxylase / Q000302:isolation & purification / Q000378:metabolism; D004298:Dopamine / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D007700:Kinetics; D008297:Male; D008970:Molecular Weight; D009532:Nickel / Q000494:pharmacology; D011732:Pyridoxal Phosphate; D051381:Rats; D012701:Serotonin / Q000494:pharmacology; D015032:Zinc / Q000494:pharmacology
D012701:Serotonin; D011732:Pyridoxal Phosphate; D004295:Dihydroxyphenylalanine; D003300:Copper; D009532:Nickel; D006916:5-Hydroxytryptophan; D004296:Dopa Decarboxylase; D015032:Zinc; D004298:Dopamine
Purification et propriétés de la L-amino acide aromatique décarboxylase (4.1.1.28) du cerveau de rat