title: Advances in structure-function relationships of tyrosinase from Agaricus bisporus - investigation on heat-induced conformational changes.
abstract: A combination of fluorescence spectroscopic measurements, inactivation kinetics and in silico prediction was used in the present study to investigate the heat induced behaviour of tyrosinase from Agaricus bisporus. The phase diagram indicated the existence of at least two distinct species induced by the temperature increase up to 75°C. Regardless of calcium ion presence, the fluorescence intensity results suggest that tyrosinase tends to form aggregates after 10min at 75°C. The quenching experiments using acrylamide and iodide demonstrate a more flexible conformation of tyrosinase at higher temperature. Detailed insights into tyrosinase structure after performing molecular dynamics simulations, suggest important structural rearrangements of the protein with the temperature increase. The copper coordinating His(94) residue was predicted to be involved in salt bridge formation with Glu(98), therefore causing significant alteration of the substrate binding site with increasing temperature. These significant changes in tyrosinase structure at temperatures over 60°C might lead to enzyme inactivation.
authors.affiliation: Dunarea de Jos University of Galati, Faculty of Food Science and Engineering, Domnească Street 111, 800201 Galati, Romania.
authors.forename: Elena
authors.initials: E
authors.lastname: Ioniţă
authors.affiliation: Dunarea de Jos University of Galati, Faculty of Food Science and Engineering, Domnească Street 111, 800201 Galati, Romania. Electronic address: iuliana.aprodu@ugal.ro.
authors.forename: Iuliana
authors.initials: I
authors.lastname: Aprodu
authors.affiliation: Dunarea de Jos University of Galati, Faculty of Food Science and Engineering, Domnească Street 111, 800201 Galati, Romania.
authors.forename: Nicoleta
authors.initials: N
authors.lastname: Stănciuc
authors.affiliation: Dunarea de Jos University of Galati, Faculty of Food Science and Engineering, Domnească Street 111, 800201 Galati, Romania.
authors.forename: Gabriela
authors.initials: G
authors.lastname: Râpeanu
authors.affiliation: Dunarea de Jos University of Galati, Faculty of Food Science and Engineering, Domnească Street 111, 800201 Galati, Romania. Electronic address: http://www.bioaliment.ugal.ro.
authors.forename: Gabriela
authors.initials: G
authors.lastname: Bahrim
mesh_terms: D000364:Agaricus / Q000737:chemistry / Q000201:enzymology*; D001665:Binding Sites; D003300:Copper / Q000378:metabolism; D004795:Enzyme Stability; D005656:Fungal Proteins / Q000737:chemistry / Q000378:metabolism; D006358:Hot Temperature; D007700:Kinetics; D014442:Monophenol Monooxygenase / Q000737:chemistry* / Q000378:metabolism*; D011487:Protein Conformation; D013050:Spectrometry, Fluorescence
chemical_list: D005656:Fungal Proteins; D003300:Copper; D014442:Monophenol Monooxygenase
keywords: Conformation; Fluorescence spectroscopy; Kinetics; Molecular dynamics; Tyrosinase