Heterologous expression and characterization of functional mushroom tyrosinase (AbPPO4)

Abstract Tyrosinases are an ubiquitous group of copper containing metalloenzymes that hydroxylate and oxidize phenolic molecules. In an application context the term ‘tyrosinase’ usually refers to ‘mushroom tyrosinase’ consisting of a mixture of isoenzymes and containing a number of enzymatic side-ac...

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Autores principales:	Matthias Pretzler, Aleksandar Bijelic, Annette Rompel
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2017
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/b7863f7a2b9a49b2aef883060dd7bef2
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spelling	oai:doaj.org-article:b7863f7a2b9a49b2aef883060dd7bef22021-12-02T16:08:12ZHeterologous expression and characterization of functional mushroom tyrosinase (AbPPO4)10.1038/s41598-017-01813-12045-2322https://doaj.org/article/b7863f7a2b9a49b2aef883060dd7bef22017-05-01T00:00:00Zhttps://doi.org/10.1038/s41598-017-01813-1https://doaj.org/toc/2045-2322Abstract Tyrosinases are an ubiquitous group of copper containing metalloenzymes that hydroxylate and oxidize phenolic molecules. In an application context the term ‘tyrosinase’ usually refers to ‘mushroom tyrosinase’ consisting of a mixture of isoenzymes and containing a number of enzymatic side-activities. We describe a protocol for the efficient heterologous production of tyrosinase 4 from Agaricus bisporus in Escherichia coli. Applying this procedure a pure preparation of a single isoform of latent tyrosinase can be achieved at a yield of 140 mg per liter of autoinducing culture medium. This recombinant protein possesses the same fold as the enzyme purified from the natural source as evidenced by single crystal X-ray diffraction. The latent enzyme can be activated by limited proteolysis with proteinase K which cleaves the polypeptide chain after K382, only one The latent enzyme can amino acid before the main in-vivo activation site. Latent tyrosinase can be used as obtained and enzymatic activity may be induced in the reaction mixture by the addition of an ionic detergent (e.g. 2 mM SDS). The proteolytically activated mushroom tyrosinase shows >50% of its maximal activity in the range of pH 5 to 10 and accepts a wide range of substrates including mono- and diphenols, flavonols and chalcones.Matthias PretzlerAleksandar BijelicAnnette RompelNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 7, Iss 1, Pp 1-10 (2017)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Matthias Pretzler Aleksandar Bijelic Annette Rompel Heterologous expression and characterization of functional mushroom tyrosinase (AbPPO4)
description	Abstract Tyrosinases are an ubiquitous group of copper containing metalloenzymes that hydroxylate and oxidize phenolic molecules. In an application context the term ‘tyrosinase’ usually refers to ‘mushroom tyrosinase’ consisting of a mixture of isoenzymes and containing a number of enzymatic side-activities. We describe a protocol for the efficient heterologous production of tyrosinase 4 from Agaricus bisporus in Escherichia coli. Applying this procedure a pure preparation of a single isoform of latent tyrosinase can be achieved at a yield of 140 mg per liter of autoinducing culture medium. This recombinant protein possesses the same fold as the enzyme purified from the natural source as evidenced by single crystal X-ray diffraction. The latent enzyme can be activated by limited proteolysis with proteinase K which cleaves the polypeptide chain after K382, only one The latent enzyme can amino acid before the main in-vivo activation site. Latent tyrosinase can be used as obtained and enzymatic activity may be induced in the reaction mixture by the addition of an ionic detergent (e.g. 2 mM SDS). The proteolytically activated mushroom tyrosinase shows >50% of its maximal activity in the range of pH 5 to 10 and accepts a wide range of substrates including mono- and diphenols, flavonols and chalcones.
format	article
author	Matthias Pretzler Aleksandar Bijelic Annette Rompel
author_facet	Matthias Pretzler Aleksandar Bijelic Annette Rompel
author_sort	Matthias Pretzler
title	Heterologous expression and characterization of functional mushroom tyrosinase (AbPPO4)
title_short	Heterologous expression and characterization of functional mushroom tyrosinase (AbPPO4)
title_full	Heterologous expression and characterization of functional mushroom tyrosinase (AbPPO4)
title_fullStr	Heterologous expression and characterization of functional mushroom tyrosinase (AbPPO4)
title_full_unstemmed	Heterologous expression and characterization of functional mushroom tyrosinase (AbPPO4)
title_sort	heterologous expression and characterization of functional mushroom tyrosinase (abppo4)
publisher	Nature Portfolio
publishDate	2017
url	https://doaj.org/article/b7863f7a2b9a49b2aef883060dd7bef2
work_keys_str_mv	AT matthiaspretzler heterologousexpressionandcharacterizationoffunctionalmushroomtyrosinaseabppo4 AT aleksandarbijelic heterologousexpressionandcharacterizationoffunctionalmushroomtyrosinaseabppo4 AT annetterompel heterologousexpressionandcharacterizationoffunctionalmushroomtyrosinaseabppo4
_version_	1718384617720905728

Heterologous expression and characterization of functional mushroom tyrosinase (AbPPO4)

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