I want to purify tyrosinase enzyme from streptomyces bacteria after production in liquid medium and I d like to use tyrosine to check the activity ?
Dear Harir,
The answer is yes. The attached file describes the use of tyrosine as a substrate for tyrosinase.
In the presence of oxygen (O2), tyrosinase (E) catalyzes the
hydroxylation of tyrosine into the compound 3,4-dihydroxyphenylalanine, or DOPA for short. Tyrosinase then catalyzes DOPA into dopaquinone which spontaneously converts into dopachrome. Dopachrome will eventually be turned into melanin. Dopachrome is a colored compound with a peak absorbance at 475 nm that can be monitored using a Spectrometer or Colorimeter.
Hoping this will be helpful,
Rafik
The above information from Rafik is correct, but I would add that the tyrosine hydroxylase activity of tyrosinase shows often a "lag" period, and the dopachrome formation is not starting from the very beginning. The lag period can be variable, from a few seconds to hours. It depends on the tyrosinase source. To avoid that, you have to add to the reaction media a catalytic amount of L-dopa (around 10-20 micromolar for around 1 mM tyrosine).
On the other hand, Streptomyces usually have protyrosinase, as this is inactive unless it would be activated. I do not know the strain that you handle, but I recomend to read some references concerning that before measuring.
Anyway, the assay using L-dopa as substrate is much easier than with L-tyrosine.
Best regards
Dear ALL,
thank you so much for your answers, I Know that L dopa is "the golden substrat " for tyrosinase enzyme , but I havn 't this substrat now and i d like to find an other substitute substrat to check the activity of my enszymes during the diffrentes step of purification .
Best regards
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