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Literature summary extracted from

  • Goldfeder, M.; Kanteev, M.; Isaschar-Ovdat, S.; Adir, N.; Fishman, A.
    Determination of tyrosinase substrate-binding modes reveals mechanistic differences between type-3 copper proteins (2014), Nat. Commun., 5, 4505 .
    View publication on PubMed

Cloned(Commentary)

EC Number Cloned (Comment) Organism
1.14.18.1 recombinant expression of His-tagged enzyme in Escherichia coli BL21 Priestia megaterium

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
1.14.18.1 purified enzyme bound with L-tyrosine or L-dopa and zinc ions, hanging drop vapour diffusion method, mixing of 0.002 ml or 2 mg/ml protein in 20mM Tris-HCl, pH 7.5, and 500 mM NaCl, with 0.002 ml reservoir solution containing 18% PEG 8000 and 0.1 M cacodylic acid, pH 6.5, and equilibration against 0.6 ml of reservoir solution, 20°C, soaking of the crystals in inhibiting ZnCl2 before soaking them in the substrate solution in order to trap the substrates within the active site of TyrBm in the crystal, X-ray diffraction structure determination and analysis at 2.2-2.5 A resolution Priestia megaterium

Inhibitors

EC Number Inhibitors Comment Organism Structure
1.14.18.1 Zn2+ binding structure in the active site, modeling, overview. Replacing the Cu2+ with Zn2+ ions can be performed in TyrBm without structural consequences, while the presence of Zn2+ ions inhibits the activity of tyrosinase on both monophenols and diphenols Priestia megaterium

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
1.14.18.1 Cu2+ the enzyme is a type-3 copper protein Priestia megaterium

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
1.14.18.1 2 L-dopa + O2 Priestia megaterium
-
 2 dopaquinone + 2 H2O
-
 ?
1.14.18.1 tyrosine + O2 Priestia megaterium
-
 dopaquinone + H2O
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
1.14.18.1 Priestia megaterium
-
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
1.14.18.1 recombinant His-tagged enzyme from Escherichia coli BL21 by nickel affinity chromatography Priestia megaterium

Reaction

EC Number Reaction Comment Organism Reaction ID
1.14.18.1 L-tyrosine + O2 = dopaquinone + H2O molecular reaction mechanism and substrate binding mode, L-tyrosine in the active site forms hydrogen bonds with R209 via its carboxyl side chain Priestia megaterium
a

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.14.18.1 2 L-dopa + O2
-
 Priestia megaterium 2 dopaquinone + 2 H2O
-
 ?
1.14.18.1 tyrosine + O2
-
 Priestia megaterium dopaquinone + H2O
-
 ?

Synonyms

EC Number Synonyms Comment Organism
1.14.18.1 TyrBm
-
 Priestia megaterium

General Information

EC Number General Information Comment Organism
1.14.18.1 metabolism tyrosinase is responsible for the two initial enzymatic steps in the conversion of tyrosine to melanin Priestia megaterium
1.14.18.1 additional information determination of tyrosinase substrate-binding modes, overview. Both monophenol hydroxylation and diphenol oxidation occur at the same site. Compared to tyrosinase, the concurrent presence of a phenylalanine above the active site and a restricting thioether bond on the histidine coordinating copper ion CuA prevent hydroxylation of monophenols by catechol oxidases, EC 1.10.3.1. A conserved water molecule activated by E195 and N205 is proposed to mediate deprotonation of the monophenol at the active site. The diphenol L-dopa binds to uinc ion ZnA in the active site in the same orientation as tyrosine, assisted by interactions with H208, both the hydroxylation and oxidation activities occur without significant binding site reorganization. Comarison of binding modes of catecol oxidase/diphenolase, and tyrosinase/monophenolase. Structure-supported monophenol hydroxylation and deprotonation mechanism, overview Priestia megaterium