Antigenicity, catalytic activity and conformation of Agaricus bisporus tyrosinase: interaction of conformation-directed antibodies with the native and irradiated enzyme.
The antiferromagnetically spin-coupled Cu2+ pair present in the active center of tyrosinase was found to be indispensable for its catalytic function. However, the metal ion did not contribute to the conformational integrity or antigenicity of the enzyme molecule. Irradiation of tyrosinase with 254 nm light resulted in dose dependent, essentially irreversible losses of its catalytic and antigenic functions. The apparent first order rate constants for the two processes were 17.6 X 10(-2) min-1 and 28.1 X 10(-2) min-1, respectively. The approximately 1.6-fold difference between the two rate constants suggests that the sites of antigenic determinants in tyrosinase are distinguishable from the enzymic active site by their higher photosensitivity. Kinetic analysis of the data as to photoinactivation, and the UV induced losses of antigenicity and structural integrity revealed that UV radiation disrupts the short-range noncovalent interactions occurring within the enzyme molecule. The disruption of the noncovalent interactions results in partial unfolding of the tyrosinase structure which in turn leads to the progressive loss of its catalytic activity and antigenicity. The anti-tyrosinase antibodies raised in rabbits were found to be directed against the native conformation of the enzyme. It is speculated that these antibodies might be useful in exploring the tyrosinase conformation and in studying the effects of various factors on the enzyme surface and molecular structure.
D000363:Agaricales / Q000201:enzymology*; D000364:Agaricus / Q000201:enzymology* / Q000276:immunology; D000818:Animals; D000908:Antibodies, Fungal; D000946:Antigens, Fungal / Q000528:radiation effects; D001665:Binding Sites; D004156:Catechol Oxidase / Q000276:immunology*; D003300:Copper; D007700:Kinetics; D014442:Monophenol Monooxygenase / Q000276:immunology* / Q000378:metabolism / Q000528:radiation effects; D011487:Protein Conformation / Q000528:radiation effects; D011817:Rabbits; D014466:Ultraviolet Rays
D000908:Antibodies, Fungal; D000946:Antigens, Fungal; D003300:Copper; D004156:Catechol Oxidase; D014442:Monophenol Monooxygenase