Małgorzata Pająk

Isotope effects in mechanistic studies of l-tyrosine halogen derivatives hydroxylation catalyzed by tyrosinase

The kinetic (KIE) and solvent (SIE) isotope effect methods were used to investigate the mechanism of enzymatic hydroxylation of halogenated derivatives of l-tyrosine to l-DOPA catalyzed by the enzyme tyrosinase (EC 1.14.18.1). The values of deuterium KIE and SIE were obtained using the non-competitive method with spectrophotometric measurements. The Lineweaver–Burk plots were used for determination of the inhibition mode of 3′-iodo-l-tyrosine. Based upon kinetic effects values the mechanism of action of enzyme tyrosinase was proposed.

Isotope effects in the hydroxylation of l -tyrosine catalyzed by tyrosinase

The reaction mechanism of the enzymatic hydroxylation of l-tyrosine (l-Tyr) to l-DOPA catalyzed by enzyme tyrosinase (EC 1.14.18.1) were investigated using the kinetic (KIE) and solvent (SIE) isotope effect methods. The values of deuterium KIE’s and SIE’s for [2′,6′-2H2]-, and [3′,5′-2H2]-l-Tyr were obtained using a non-competitive spectrophotometric method but values of H/T and D/T KIE’s for the 3′- and 5′-positions of [3′,5′-3H2]-, and [3′,5′-2H/3H]-l-Tyr were determined by competitive method with [1-14C]-l-Tyr as internal radioactive standard.

Isotope effects in the tyrosinase catalysed hydroxylation of l-tyrosine methyl derivatives

ABSTRACT To investigate isotope effects in the hydroxylation of [3′,5′-2H2]-α-methyl- and [3′,5′-2H2]-N-methyl-l-tyrosine, they were synthesised using acid catalysed isotope exchange at high temperature. The kinetic and solvent deuterium isotope effects on Vmax and Vmax/Km parameters of tyrosinase in its action on methylated derivatives of l-tyrosine were determined using the non-competitive spectrophotometric method. Lineweaver–Burk plots were used to consider the inhibition type of O-methyl-l-tyrosine, revealing that it is an uncompetitive inhibitor of tyrosinase.