Substrate Construes the Copper and Nickel Ions Impacts on the Mushroom Tyrosinase Activities

Abstract

Mushroom tyrosinase (MT) structural changes in the presence of <TEX>$Cu ^{2+}$</TEX> and <TEX>$Ni ^{2+}$</TEX> were studied separately. Far-UV CD spectra of the incubated MT with the either of the metal ions indicated reduction of the well-ordered secondary structure of the enzyme. Increasing in the maximum fluorescence emission of anilinonaphthalene-8-sulfonic acid (ANS) was also revealing partial unfolding caused by the conformational changes in the tertiary structure of MT. Thermodynamic studies on the chemical denaturation of MT by dodecyl trimethylammonium bromide (DTAB) showed decrease in the stability of MT in the presence of <TEX>$Cu ^{2+}$</TEX> or <TEX>$Ni ^{2+}$</TEX> using their activation concentrations. Both activities of MT were also assessed in the presence of different concentrations of these ions, separately, with various monophenols and their corresponding diphenols. Kinetic studies revealed that cresolase activity on p-coumaric acid was boosted in the presence of either of the metal ions, but inhibited when phenol, L-tyrosine, or 4-[(4-methylphenyl)azo]-phenol was substrate. Similarly, catecholase activity on caffeic acid was enhanced in the presence of <TEX>$Cu ^{2+}$</TEX> or <TEX>$Ni ^{2+}$</TEX>, but inhibited when catechol, L-DOPA, or 4-[(4-methylbenzo)azo]-1,2-benzenediol was substrate. Results of this study suggest that both cations make MT more fragile and less active. However, the effect of the substrate structure on the MT allosteric behavior can not be ignored.