Aggregation equilibria of tyrosinase of Harding-Passey mouse melanoma

García-Borrón, J.C.; Solano, Francisco; Iborra, J. L.; Lozano, J A

doi:10.1042/bj2280095

Cited by 12 publications

(4 citation statements)

References 26 publications

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“…Third, the LEMT and HEMT appear to have an autoaggregation behaviour that might account for their co-purification by most chromatographic techniques. If these forms actually correspond to the b and c proteins, this is not a surprising observation since purified tyrosinase is known to display autoaggregation [36] and both the b and c proteins share sufficient sequence similarity to expect the formation of heterodimers. Moreover, the presence of the b and c proteins together with other melanosomal proteins in large complexes, has also been reported 1371.…”

Section: Discussionmentioning

confidence: 99%

Tyrosinase isoenzymes in mammalian melanocytes

Jiménez‐Cervantes

García‐Borrón

Valverde

et al. 1993

European Journal of Biochemistry

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B-16 mouse melanoma melanosomes contain two forms of tyrosinase that can be resolved by SDSPAGE. These forms interact to different extents with the ion-exchanger DEAE-Sephadex and with hydroxyapatite, and have different affinity for the melanosomal membrane andor the intraorganular matrix. After partial purification and complete separation of the two tyrosinases, several kinetic parameters were analyzed. The form of lower electrophoretic mobility displayed a higher Ktn for 3,4-dihydroxy-~-phenylalanine (L-dopa) and L-tyrosine, an absolute requirement for the cofactor L-dopa in its tyrosine hydroxylase activity, and a lower ratio of tyrosine hydroxylation to Dopa oxidation. The form of higher electrophoretic mobility displayed lower values of K,, for both substrates and was able to exhibit tyrosine hydroxylase activity after a lag period even in the absence of L-dopa. Both forms were stereospecific for the L isomers and sensitive to the specific tyrosinase inhibitor 2-phenylthiourea. These forms do not appear to result from different degrees of glycosylation, nor from limited proteolysis and are also present in the microsomal fraction of B16 mouse melanoma. They might correspond to different gene products, most likely derived from the b and c loci.Mammalian tyrosinase (monophenol monooxygenase) is a copper-containing glycoprotein responsible for the synthesis of melanin within melanocytes [l, 21. The enzyme catalyses the rate-limiting step in melanin biosynthesis, the hydroxylation of L-tyrosine to 3,4-dihydroxy-~-phenylalanine @-dopa) and the subsequent oxidation of L-dopa to L-dopaquinone. In the absence of thiol compounds L-dopaquinone undergoes a rapid oxidation and spontaneous rearrangement leading to L-dopachrome, and ultimately, to the melanin polymer [3]. Although melanin formation can proceed spontaneously from L-dopaquinone, at least one other enzyme involved in the process has been characterized [4-61. This enzyme, called dopachrome tautomerase [5], catalyses the non-decarboxylative rearrangement of L-dopachrome, yielding 5,6-dihydroxyindole 2-carboxylic acid. In the absence of the enzyme, L-dopachrome spontaneously evolves to 5,6-dihydroxyindole. The series of oxidation and polymerization reactions leading from 5,6-dihydroxyindole 2-carboxylic acid and 5,6-dihydroxyindole to melanin is still poorly understood, although it has been shown that 5,6-dihydroxyindole is essential for the in vitro polymerization of its 2-carboxylic acid [7], and that tyrosinase might play a role in this distal phase of melanogenesis [7, 81.

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Section: Discussionmentioning

confidence: 99%

Tyrosinase isoenzymes in mammalian melanocytes

Jiménez‐Cervantes

García‐Borrón

Valverde

et al. 1993

European Journal of Biochemistry

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“…Because tyrosinase is an integral membrane protein, the melanosomal preparation must be detergent‐solubilized (Jiménez‐Cervantes et al., ) or a soluble, enzymatically active fragment can be obtained by limited proteolysis (Valverde et al., ). The resulting extracts can be fractionated by combinations of techniques such as ammonium sulfate precipitation, gel filtration or ion‐exchange chromatography, and preparative electrophoresis to obtain highly purified tyrosinase preparations (Garcia‐Borron et al., ; Hearing et al., ; Ohkura et al., ; Tomita et al., ). However, the different purification protocols described to date are time‐consuming, and their yields are relatively low.…”

Section: Enzymes For In Vitro Studiesmentioning

confidence: 99%

Melanins and melanogenesis: methods, standards, protocols

d’Ischia

Wakamatsu

Napolitano

et al. 2013

Pigment Cell Melanoma Res

400

482

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Subscribe to PCMR and stay up-to-date with the only journal committed to publishing basic research in melanoma and pigment cell biology As a member of the IFPCS or the SMR you automatically get online access to PCMR. Sign up as a member today at www.ifpcs.org or at www.societymelanomaresarch.org SummaryDespite considerable advances in the past decade, melanin research still suffers from the lack of universally accepted and shared nomenclature, methodologies, and structural models. This paper stems from the joint efforts of chemists, biochemists, physicists, biologists, and physicians with recognized and consolidated expertise in the field of melanins and melanogenesis, who critically reviewed and experimentally revisited methods, standards, and protocols to provide for the first time a consensus set of recommended procedures to be adopted and shared by researchers involved in pigment cell research. The aim of the paper was to define an unprecedented frame of reference built on cutting-edge knowledge and state-of-the-art methodology, to enable reliable comparison of results among laboratories and new progress in the field based on standardized methods and shared information.

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“…Optical measurements of natural melanins are largely based on their absorption properties (e.g., [10,14,15]), but this has several limitations because melanins have no distinctive absorption peaks that allow distinguishing them from other pigments. Toral et al [10] made discriminant analyses that allowed the separation of melanins and carotenoids in feathers based on the shape of their reflectance spectra, but also highlighted the difficulty in distinguishing between pheomelanin and other pigments with similar reflectance profiles such as iron oxide and porphyrins.…”

Section: Introductionmentioning

confidence: 99%