Changes in tyrosinase specificity by ionic liquids and sodium dodecyl sulfate
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Tyrosinase is a member of the type 3 copper enzyme family involved in the production of melanin in a wide range of organisms. The ability of tyrosinases to convert monophenols into diphenols has stimulated studies regarding the production of substituted catechols, important intermediates for the synthesis of pharmaceuticals, agrochemicals, polymerization inhibitors, and antioxidants. Despite its enormous potential, the use of tyrosinases for catechol synthesis has been limited due to the low monophenolase/diphenolase activity ratio. In the presence of two water miscible ionic liquids, [BMIM][BF4] and ethylammonium nitrate, the selectivity of a tyrosinase from Bacillus megaterium (TyrBm) was altered, and the ratio of monophenolase/diphenolase activity increased by up to 5-fold. Furthermore, the addition of sodium dodecyl sulphate (SDS) at levels of 2–50 mM increased the activity of TyrBm by 2-fold towards the natural substrates l-tyrosine and l-Dopa and 15- to 20-fold towards the non-native phenol and catechol. The R209H tyrosinase variant we previously identified as having a preferential ratio of monophenolase/diphenolase activity was shown to have a 45-fold increase in activity towards phenol in the presence of SDS. We propose that the effect of SDS on the ability of tyrosinase to convert non-natural substrates is due to the interaction of surfactant molecules with residues located at the entrance to the active site, as visualized by the newly determined crystal structure of TyrBm in the presence of SDS. The effect of SDS on R209 may enable less polar substrates such as phenol and catechol, to penetrate more efficiently into the enzyme catalytic pocket.
KeywordsTyrosinase Bacillus megaterium Ionic liquids Sodium dodecyl sulphate Diphenols
This work was supported by the Israel Science Foundation founded by the Israel Academy of Sciences and Humanities, grant number 193/11. We gratefully thank the staff of the ESRF (beamline ID23-1) for provision of synchrotron radiation facilities and assistance.
- Adams PD, Afonine PV, Bunkoczi G, Chen VB, Davis IW, Echols N, Headd JJ, Hung LW, Kapral GJ, Grosse-Kunstleve RW, McCoy AJ, Moriarty NW, Oeffner R, Read RJ, Richardson DC, Richardson JS, Terwilliger TC, Zwart PH (2010) PHENIX: a comprehensive Python-based system for macromolecular structure solution. Acta Crystallogr Sect D Biol Crystallogr 66: 213–221.CrossRefGoogle Scholar
- Brünger AT, Adams PD, Clore GM, DeLano WL, Gros P, Grosse-Kunstleve RW, Jiang JS, Kuszewski J, Nilges M, Pannu NS, Read RJ, Rice LM, Simonson T, Warren GL (1998) Crystallography & NMR system: A new software suite for macromolecular structure determination. Acta Crystallogr Sect D Biol Crystallogr 54: 905–921.CrossRefGoogle Scholar
- Karbassi F, Haghbeen K, Saboury AA, Rezaei-Tavirani M, Ranjbar B (2004) Calorimetric, spectrophotometric and circular dichroism studies on the impact of sodium dodecyl sulfate on the mushroom tyrosinase structure. Biologia 59: 319–326.Google Scholar
- Leslie AGW (1992) Joint CCP4 + ESF-EAMCB Newsletter on protein crystallography: No. 26.Google Scholar
- Moore BM, Flurkey WH (1990) Sodium dodecyl sulfate activation of a plant polyphenoloxidase. Effect of sodium dodecyl sulfate on enzymatic and physical characteristics of purified broad bean polyphenoloxidase. J Biol Chem 265: 4982–4988.Google Scholar