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Literature summary for 1.14.18.1 extracted from

  • Dolinska, M.B.; Wingfield, P.T.; Sergeev, Y.V.
    Purification of recombinant human tyrosinase from insect larvae infected with the baculovirus vector (2017), Curr. Protoc. Protein Sci., 89, 15.1-6.15.12 .
    View publication on PubMedView publication on EuropePMC
Search for more literature for 1.14.18.1

Cloned(Commentary)

Cloned (Comment) Organism
gene TYR, DNA and amino acid sequence determinatin and analysis, sequence comparisons, recombinant expression of His-tagged intra-melanosomal domain wild-type and mutant variants OCA1A and OCA1B in Trichoplusia ni insect cells using the baculovirus transfection method Homo sapiens
recombinant expression of His-tagged intra-melanosomal domain of wild-type tyrosinase and temperature-sensitive OCA1-related mutant R422Q and in whole-insect Trichoplusia ni via transfection with baculovius Homo sapiens

Protein Variants

Protein Variants Comment Organism
additional information generation of the recombinant human tyrosinase intra-melanosomal domain and mutant variants, which mimic genetic changes in both subtypes of OCA1 patients, by site-directed mutagenesis and recombinant expression in insect cells. The recombinant OCA1A show very low protein expression, protein yield, and are enzymatically inactive, while mutants mimicking OCA1B are biochemically similar to the wild-type, but exhibit lower specific activities and protein stabilities than the wild-type enzyme Homo sapiens
additional information the strategy of deleting the membrane-anchoring helix is standard to simplify handling of protein. Deletion of the anchoring helix of the human tyrosinase does not affect the tyrosinase activity Homo sapiens
P406L a phenotype OCA1A-related mutant, inactive mutant Homo sapiens
R402Q a phenotype OCA1A-related mutant, inactive mutant Homo sapiens
R422Q a phenotype OCA1A-related mutant, inactive mutant Homo sapiens
R422W a phenotype OCA1A-related mutant, inactive mutant Homo sapiens
R77Q a phenotype OCA1A-related mutant, inactive mutant Homo sapiens
T373K a phenotype OCA1A-related mutant, inactive mutant Homo sapiens

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
 additional information Michaelis-Menten kinetics Homo sapiens

Localization

Localization Comment Organism GeneOntology No. Textmining
melanosome
-
 Homo sapiens 42470
-
membrane membrane associated enzyme, deletion of the anchoring helix does not affect the tyrosinase activity Homo sapiens 16020
-
membrane tyrosinase is a membrane glycoprotein Homo sapiens 16020
-

Metals/Ions

Metals/Ions Comment Organism Structure
Cu2+ a copper-containing enzyme, the two copper ions (CuA and CuB) essential for activity Homo sapiens

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
2 L-dopa + O2 Homo sapiens
-
 2 dopaquinone + 2 H2O
-
 ?
tyrosine + O2 Homo sapiens
-
 dopaquinone + H2O
-
 ?

Organism

Organism UniProt Comment Textmining
Homo sapiens P14679
-
-

Posttranslational Modification

Posttranslational Modification Comment Organism
glycoprotein tyrosinase is a membrane glycoprotein Homo sapiens
glycoprotein tyrosinase is a membrane-associated glycoprotein. The purified recombinant human tyrosinase is a soluble monomeric glycoprotein. N-Glycosylation of tyrosinase appears critical for enzyme stability and enzymatic function Homo sapiens

Purification (Commentary)

Purification (Comment) Organism
recombinant His-tagged intra-melanosomal domain of wild-type tyrosinase and temperature-sensitive OCA1-related mutant R422Q from whole-insect Trichoplusia ni by nickel affinity chromatography, tag cleavage by TEV protease, dialysis, and two steps of gel filtration, 98.73% purity. The purified recombinant human tyrosinase is a soluble monomeric glycoprotein with an activity that mirrors the tyrosinase in vivo function Homo sapiens
recombinant His-tagged intra-melanosomal domain wild-type and mutant variants OCA1A and OCA1B in Trichoplusia ni insect cells by nickel affinity chromatography, ultrafiltration, and gel filtration Homo sapiens

Renatured (Commentary)

Renatured (Comment) Organism
all proteins including hTyrCtr and OCA1B-related mutant variants are a subjected to equilibrium unfolding/refolding using 0.001 mM protein in 10 mM phosphate buffer, pH 7.4, 5 h at 8 M urea, followed by dilution with 10 mM phosphate buffer, pH 7.4 to 0-8 M urea and incubated for 24 h at room temperature Homo sapiens

Source Tissue

Source Tissue Comment Organism Textmining

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
350.02
-
 purified recombinant wild-type intra-melanosomal domain, pH 7.4, 37°C Homo sapiens

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2 L-dopa + O2
-
 Homo sapiens 2 dopaquinone + 2 H2O
-
 ?
tyrosine + O2
-
 Homo sapiens dopaquinone + H2O
-
 ?

Subunits

Subunits Comment Organism
? x * 56630, recombinant enzyme, SDS-PAGE Homo sapiens
monomer x * 57000, recombinant enzyme, SDS-PAGE Homo sapiens

Synonyms

Synonyms Comment Organism
catecholase
-
 Homo sapiens
cresolase
-
 Homo sapiens
Diphenol oxidase
-
 Homo sapiens
L-DOPA monophenolase
-
 Homo sapiens
OCA1
-
 Homo sapiens
tyr
-
 Homo sapiens

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
37
-
 assay at Homo sapiens

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.4
-
 assay at Homo sapiens

General Information

General Information Comment Organism
malfunction mutations in the tyrosinase gene cause oculocutaneous albinism type 1 (OCA1), an autosomal recessive disease associated with reduced melanin pigment in the hair, skin, and eyes and decreased quality of vision Homo sapiens
malfunction oculocutaneous albinism Type 1 (OCA1) is an autosomal recessive disorder caused by mutations in the tyrosinase gene. Two subtypes of OCA1 exxist, severe OCA1A with complete absence of tyrosinase activity and less severe OCA1B with residual tyrosinase activity. The recombinant OCA1A mutants expressed in insect cells show very low protein expression, protein yield, and are enzymatically inactive, while mutants mimicking OCA1B are biochemically similar to the wild-type, but exhibit lower specific activities and protein stabilities than the wild-type enzyme. OCA1A mutations inactivate tyrosinase and result in severe phenotype, while OCA1B mutations partially inactive tyrosinase and results in OCA1B albinism Homo sapiens
metabolism tyrosinase catalyzes the first two steps of the melanin synthesis pathway: hydroxylation of L-tyrosine to L-3,4-dihydroxyphenylalanine, L-DOPA, monophenolase or cresolase activity, EC1.14.18.1, and the subsequent oxidation of L-DOPA to dopaquinone, diphenol oxidase or catecholase activity, EC 1.10.3.1 Homo sapiens
additional information homology model of human tyrosinase incorporated into the phospholipid membrane, overview Homo sapiens
additional information structure homology modeling of wild-type an dmutant enzymes Homo sapiens
physiological function the enzyme catalyzes the oxidation of L-tyrosine (monophenol substrate) and L-3,4-dihydroxyphenylalanine (L-DOPA, diphenol substrate) to form dopaquinone. The monophenolase and diphenol oxidase activities are linked to the tyrosinase active site, which is composed of six histidine residues that coordinate two copper ions (CuA and CuB) essential for activity Homo sapiens