pH-induced structural changes of tyrosinase from Agaricus bisporus using fluorescence and in silico methods

Tyrosinases are involved in enzymatic browning reactions in damaged fruits during post-harvest handling and processing. The overall structure of tyrosinase from Agaricus bisporus mushrooms at different pH values was monitored using fluorescence spectroscopy and molecular dynamics simulations.

When the pH value was increased from 6.0 to 9.0, the protein passed through several structural intermediates, including the tetramer, trimer and dimer stages. Changes in the secondary and tertiary structure of tyrosinase at neutral pH were outlined after running molecular dynamics simulations. A detailed check at the single-molecule level by means of molecular modeling tools suggested that the most important contribution to the fluorescence intensity is given by the H subunits with seven Trp and nine Tyr residues exposed to the solvent, whereas the lectin-like folded L subunits have only six Trp and three Tyr residues, of which only Trp(15) , Trp(59) and Trp(93) are partially exposed to the solvent.

The results indicated that the enzyme was sensitive to pH. The experimental results revealed the unfolding of the native tetrameric enzyme in acidic pH range, causing exposure of the hydrophobic residues.