Kinetics of the slow pH-mediated transition of polyphenol oxidase.
Catecholase activity of latent polyphenol oxidase from broad bean leaves showed a hysteresis phenomenon above pH 4, whereas a steady-state rate was reached immediately when pH values were lower, thus suggesting that slow pH-induced conformational changes in the protein occur during the assay. When the enzyme was activated by sodium dodecyl sulfate, the lag period completely disappeared. This transition was reversible, since a burst was observed when the enzyme was preincubated at acid pH, before being returned to its previous experimental conditions. The pK for the isomerization process (pK(H) = 4.6) was estimated by preincubating the enzyme at different pH values and analyzing the product accumulation curves. Negative kinetic cooperativity was evident over a pH range in which the isomerization reaction was significant when the steady state was measured as a function of different substrate concentrations.
Jiménez
M
M
Departamento de Bioquímica y Biología Molecular A, Facultad de Biología, Universidad de Murcia, Campus de Espinardo, Spain.
García-Carmona
F
F
D004156:Catechol Oxidase / Q000737:chemistry / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects; D007887:Fabaceae / Q000201:enzymology; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D018515:Plant Leaves / Q000201:enzymology; D010946:Plants, Medicinal; D011487:Protein Conformation; D012967:Sodium Dodecyl Sulfate / Q000494:pharmacology
D012967:Sodium Dodecyl Sulfate; D004156:Catechol Oxidase