Three recombinantly expressed apple tyrosinases suggest the amino acids responsible for mono- versus diphenolase activity in plant polyphenol oxidases

Abstract Tyrosinases and catechol oxidases belong to the polyphenol oxidase (PPO) enzyme family, which is mainly responsible for the browning of fruits. Three cDNAs encoding PPO pro-enzymes have been cloned from leaves of Malus domestica (apple, MdPPO). The three pro-enzymes MdPPO1-3 were heterologo...

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Autores principales:	Ioannis Kampatsikas, Aleksandar Bijelic, Matthias Pretzler, Annette Rompel
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2017
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/0eb697f1a73343dead73f6f62c3ee7ff
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spelling	oai:doaj.org-article:0eb697f1a73343dead73f6f62c3ee7ff2021-12-02T12:30:18ZThree recombinantly expressed apple tyrosinases suggest the amino acids responsible for mono- versus diphenolase activity in plant polyphenol oxidases10.1038/s41598-017-08097-52045-2322https://doaj.org/article/0eb697f1a73343dead73f6f62c3ee7ff2017-08-01T00:00:00Zhttps://doi.org/10.1038/s41598-017-08097-5https://doaj.org/toc/2045-2322Abstract Tyrosinases and catechol oxidases belong to the polyphenol oxidase (PPO) enzyme family, which is mainly responsible for the browning of fruits. Three cDNAs encoding PPO pro-enzymes have been cloned from leaves of Malus domestica (apple, MdPPO). The three pro-enzymes MdPPO1-3 were heterologously expressed in E. coli yielding substantial amounts of protein and have been characterized with regard to their optimum of activity resulting from SDS, acidic and proteolytic activation. Significant differences were found in the kinetic characterization of MdPPO1-3 when applying different mono- and diphenolic substrates. All three enzymes have been classified as tyrosinases, where MdPPO1 exhibits the highest activity with tyramine (kcat = 9.5 s−1) while MdPPO2 and MdPPO3 are also clearly active on this monophenolic substrate (kcat = 0.92 s−1 and kcat = 1.0 s−1, respectively). Based on the activity, sequence data and homology modelling it is proposed that the monophenolase and diphenolase activity of PPOs can be manipulated by the appropriate combination of two amino acids, which are located within the active site cleft and were therefore named “activity controllers”.Ioannis KampatsikasAleksandar BijelicMatthias PretzlerAnnette RompelNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 7, Iss 1, Pp 1-13 (2017)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Ioannis Kampatsikas Aleksandar Bijelic Matthias Pretzler Annette Rompel Three recombinantly expressed apple tyrosinases suggest the amino acids responsible for mono- versus diphenolase activity in plant polyphenol oxidases
description	Abstract Tyrosinases and catechol oxidases belong to the polyphenol oxidase (PPO) enzyme family, which is mainly responsible for the browning of fruits. Three cDNAs encoding PPO pro-enzymes have been cloned from leaves of Malus domestica (apple, MdPPO). The three pro-enzymes MdPPO1-3 were heterologously expressed in E. coli yielding substantial amounts of protein and have been characterized with regard to their optimum of activity resulting from SDS, acidic and proteolytic activation. Significant differences were found in the kinetic characterization of MdPPO1-3 when applying different mono- and diphenolic substrates. All three enzymes have been classified as tyrosinases, where MdPPO1 exhibits the highest activity with tyramine (kcat = 9.5 s−1) while MdPPO2 and MdPPO3 are also clearly active on this monophenolic substrate (kcat = 0.92 s−1 and kcat = 1.0 s−1, respectively). Based on the activity, sequence data and homology modelling it is proposed that the monophenolase and diphenolase activity of PPOs can be manipulated by the appropriate combination of two amino acids, which are located within the active site cleft and were therefore named “activity controllers”.
format	article
author	Ioannis Kampatsikas Aleksandar Bijelic Matthias Pretzler Annette Rompel
author_facet	Ioannis Kampatsikas Aleksandar Bijelic Matthias Pretzler Annette Rompel
author_sort	Ioannis Kampatsikas
title	Three recombinantly expressed apple tyrosinases suggest the amino acids responsible for mono- versus diphenolase activity in plant polyphenol oxidases
title_short	Three recombinantly expressed apple tyrosinases suggest the amino acids responsible for mono- versus diphenolase activity in plant polyphenol oxidases
title_full	Three recombinantly expressed apple tyrosinases suggest the amino acids responsible for mono- versus diphenolase activity in plant polyphenol oxidases
title_fullStr	Three recombinantly expressed apple tyrosinases suggest the amino acids responsible for mono- versus diphenolase activity in plant polyphenol oxidases
title_full_unstemmed	Three recombinantly expressed apple tyrosinases suggest the amino acids responsible for mono- versus diphenolase activity in plant polyphenol oxidases
title_sort	three recombinantly expressed apple tyrosinases suggest the amino acids responsible for mono- versus diphenolase activity in plant polyphenol oxidases
publisher	Nature Portfolio
publishDate	2017
url	https://doaj.org/article/0eb697f1a73343dead73f6f62c3ee7ff
work_keys_str_mv	AT ioanniskampatsikas threerecombinantlyexpressedappletyrosinasessuggesttheaminoacidsresponsibleformonoversusdiphenolaseactivityinplantpolyphenoloxidases AT aleksandarbijelic threerecombinantlyexpressedappletyrosinasessuggesttheaminoacidsresponsibleformonoversusdiphenolaseactivityinplantpolyphenoloxidases AT matthiaspretzler threerecombinantlyexpressedappletyrosinasessuggesttheaminoacidsresponsibleformonoversusdiphenolaseactivityinplantpolyphenoloxidases AT annetterompel threerecombinantlyexpressedappletyrosinasessuggesttheaminoacidsresponsibleformonoversusdiphenolaseactivityinplantpolyphenoloxidases
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Three recombinantly expressed apple tyrosinases suggest the amino acids responsible for mono- versus diphenolase activity in plant polyphenol oxidases

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