Determination of phenolic compounds using recombinant tyrosinase from Streptomyces antibioticus

Properties of Streptomyces antibioticus tyrosinase and the implementation of the enzyme in a biosensor for the detection of phenolic compounds were investigated. The tyrosinase from S. antibioticus is a monomer and has a molecular weight of 30.6kD. The specific activity is about 5U/mg with catechol as substrate and 1225U/mg with L-dopa as substrate. The activity of tyrosinase upon long-term storage is best maintained in buffer at temperatures of -80 or + 4°C. Storage at -18°C, with or without glycerol, resulted in quick enzyme inactivation. For the construction of the sensor bi-enzymatic substrate recycling was exploited. Quinoprotein glucose dehydrogenase (GDH) and tyrosinase were immobilised in polyvinyl alcohol and coupled to a Clark-type oxygen electrode that allowed for monitoring of the oxygen consumption during catechol conversion. This design of the sensor facilitates the determination of phenolic compounds in the nanomolar range. The lower limit of detection for L-dopa, dopamine, and adrenalin was 5nM.