RESUMO
The reaction between mushroom tyrosinase and L-ascorbic acid was studied by oxymetric assays and evidence pointing to ascorbate oxidase activity of this enzyme has been obtained. The activity is clearly linear to enzyme concentration and the Michaelis constant for L-ascorbic acid has a value of 2.69 +/- 0.11 mM. Maximum activity is obtained at pH 7.5. A possible reaction mechanism, which is based on the different enzymatic forms of tyrosinase, is also presented.
Assuntos
Ascorbato Oxidase/metabolismo , Ácido Ascórbico/metabolismo , Basidiomycota/enzimologia , Monofenol Mono-Oxigenase/metabolismo , Cloretos/farmacologia , Concentração de Íons de Hidrogênio , Cinética , Melaninas/biossíntese , Estrutura Molecular , Monofenol Mono-Oxigenase/antagonistas & inibidores , Oxigênio/metabolismo , Tropolona/farmacologiaRESUMO
A kinetic analysis of the glutamine synthetase bicyclic cascade is presented. It includes the dependence on time from the onset of the reaction of both the uridylylation of Shapiro's regulatory protein and the adenylylation of the glutamine synthetase. The transient phase equations obtained allow an estimation of the time elapsed until the states of uridylylation and adenylylation reach their steady-states, and therefore an evaluation of the effective sensitivity of the system. The contribution of the uridylylation cycle to the adenylylation cycle has been studied, and an equation relating the state of adenylylation at any time to the state of uridylylation at the same instant has been derived.
Assuntos
Glutamato-Amônia Ligase/metabolismo , Trifosfato de Adenosina/metabolismo , Glutamato-Amônia Ligase/química , Cinética , Periodicidade , Uridina Trifosfato/metabolismoRESUMO
A kinetic analysis of the Michaelis-Menten mechanism is made for the cases in which the free enzyme, or the enzyme-substrate complex, or both, are unstable, either spontaneously or as a result of the addition of a reagent. The explicit time-course equations of all of the species involved has been derived under conditions of limiting enzyme concentration. The validity of these equations has been checked by using numerical simulations. An experimental design and a kinetic data analysis allowing the evaluation of the parameters and kinetic constants are recommended.
Assuntos
Estabilidade Enzimática , Enzimas/metabolismo , Catálise , Simulação por Computador , Inibidores Enzimáticos/farmacologia , Cinética , Matemática , Análise de RegressãoRESUMO
Tyrosinase usually catalyzes the conversion of monophenols to o-diphenols and the oxidation of o-diphenols to the corresponding quinones. However, when 3,4-dihydroxymandelic acid was provided as the substrate, 3,4-dihydroxybenzaldehyde was produced. These results led to the proposal that tyrosinase catalyzes an unusual oxidative decarboxylation of this substrate (Sugumaran, M. (1986) Biochemistry 25, 4489-4492). However, 3,4-dihydroxybenzaldehyde is also obtained through the oxidation of 3,4-dihydroxymandelic acid by sodium periodate and on a mercury electrode. These results led to the proposal that tyrosinase catalyzes the oxidation of the substrate into o-quinone, which reacts immediately with a molecule of substrate, oxidizing it and through decarboxylation generates an intermediate (quinone methide) which transforms into 3,4-dihydroxybenzaldehyde; simultaneously, the original o-quinone is reduced to 3,4-dihydroxymandelic acid.
