Experimental and bioinformatic investigation of the proteolytic degradation of the C-terminal domain of a fungal tyrosinase

G. Faccio (Corresponding Author), Mikko Arvas, L. Thöny-Meyer, Markku Saloheimo

Research output: Contribution to journalArticleScientificpeer-review

10 Citations (Scopus)

Abstract

Proteolytic processing is a key step in the production of polyphenol oxidases such as tyrosinases, converting the inactive proenzyme to an active form. In general, the fungal tyrosinase gene codes for a ~ 60 kDa protein that is, however, isolated as an active enzyme of ~ 40 kDa, lacking the C-terminal domain. Using the secreted tyrosinase 2 from Trichoderma reesei as a model protein, we performed a mutagenesis study of the residues in proximity of the experimentally determined cleavage site which are possibly involved in the proteolytic process. However, the mutant forms of tyrosinase 2 were not secreted in a full-length form retaining the C-terminal domain, but they were processed to give a ~ 45 kDa active form. Aiming at explaining this phenomenon, we analysed in silico the properties of the C-terminal domain of tyrosinase 2, of 23 previously retrieved homologous tyrosinase sequences from fungi (C. Gasparetti, G. Faccio, M. Arvas, J. Buchert, M. Saloheimo, K. Kruus, Appl. Microbiol. Biotechnol. 86 (2010) 213–226) and of nine well-characterised polyphenol oxidases. Based on the results of our study, we exclude the key role of specific amino acids at the cleavage site in the proteolytic process and report an overall higher sensitivity to proteolysis of the linker region and of the whole C-terminal domain of fungal tyrosinases.
Original languageEnglish
Pages (from-to)37-45
Number of pages9
JournalJournal of Inorganic Biochemistry
Volume121
DOIs
Publication statusPublished - 2013
MoE publication typeA1 Journal article-refereed

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Monophenol Monooxygenase
Bioinformatics
Computational Biology
Degradation
Catechol Oxidase
Fungal Genes
Proteolysis
Mutagenesis
Enzyme Precursors
Trichoderma
Sequence Homology
Fungi
Computer Simulation
Proteins
Genes
Amino Acids
Enzymes
Processing

Keywords

  • c-terminal domain
  • fungal tyrosinase
  • protein processing
  • proteolytic activation
  • sensitivity to proteolysis

Cite this

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title = "Experimental and bioinformatic investigation of the proteolytic degradation of the C-terminal domain of a fungal tyrosinase",
abstract = "Proteolytic processing is a key step in the production of polyphenol oxidases such as tyrosinases, converting the inactive proenzyme to an active form. In general, the fungal tyrosinase gene codes for a ~ 60 kDa protein that is, however, isolated as an active enzyme of ~ 40 kDa, lacking the C-terminal domain. Using the secreted tyrosinase 2 from Trichoderma reesei as a model protein, we performed a mutagenesis study of the residues in proximity of the experimentally determined cleavage site which are possibly involved in the proteolytic process. However, the mutant forms of tyrosinase 2 were not secreted in a full-length form retaining the C-terminal domain, but they were processed to give a ~ 45 kDa active form. Aiming at explaining this phenomenon, we analysed in silico the properties of the C-terminal domain of tyrosinase 2, of 23 previously retrieved homologous tyrosinase sequences from fungi (C. Gasparetti, G. Faccio, M. Arvas, J. Buchert, M. Saloheimo, K. Kruus, Appl. Microbiol. Biotechnol. 86 (2010) 213–226) and of nine well-characterised polyphenol oxidases. Based on the results of our study, we exclude the key role of specific amino acids at the cleavage site in the proteolytic process and report an overall higher sensitivity to proteolysis of the linker region and of the whole C-terminal domain of fungal tyrosinases.",
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author = "G. Faccio and Mikko Arvas and L. Th{\"o}ny-Meyer and Markku Saloheimo",
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Experimental and bioinformatic investigation of the proteolytic degradation of the C-terminal domain of a fungal tyrosinase. / Faccio, G. (Corresponding Author); Arvas, Mikko; Thöny-Meyer, L.; Saloheimo, Markku.

In: Journal of Inorganic Biochemistry, Vol. 121, 2013, p. 37-45.

Research output: Contribution to journalArticleScientificpeer-review

TY - JOUR

T1 - Experimental and bioinformatic investigation of the proteolytic degradation of the C-terminal domain of a fungal tyrosinase

AU - Faccio, G.

AU - Arvas, Mikko

AU - Thöny-Meyer, L.

AU - Saloheimo, Markku

PY - 2013

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N2 - Proteolytic processing is a key step in the production of polyphenol oxidases such as tyrosinases, converting the inactive proenzyme to an active form. In general, the fungal tyrosinase gene codes for a ~ 60 kDa protein that is, however, isolated as an active enzyme of ~ 40 kDa, lacking the C-terminal domain. Using the secreted tyrosinase 2 from Trichoderma reesei as a model protein, we performed a mutagenesis study of the residues in proximity of the experimentally determined cleavage site which are possibly involved in the proteolytic process. However, the mutant forms of tyrosinase 2 were not secreted in a full-length form retaining the C-terminal domain, but they were processed to give a ~ 45 kDa active form. Aiming at explaining this phenomenon, we analysed in silico the properties of the C-terminal domain of tyrosinase 2, of 23 previously retrieved homologous tyrosinase sequences from fungi (C. Gasparetti, G. Faccio, M. Arvas, J. Buchert, M. Saloheimo, K. Kruus, Appl. Microbiol. Biotechnol. 86 (2010) 213–226) and of nine well-characterised polyphenol oxidases. Based on the results of our study, we exclude the key role of specific amino acids at the cleavage site in the proteolytic process and report an overall higher sensitivity to proteolysis of the linker region and of the whole C-terminal domain of fungal tyrosinases.

AB - Proteolytic processing is a key step in the production of polyphenol oxidases such as tyrosinases, converting the inactive proenzyme to an active form. In general, the fungal tyrosinase gene codes for a ~ 60 kDa protein that is, however, isolated as an active enzyme of ~ 40 kDa, lacking the C-terminal domain. Using the secreted tyrosinase 2 from Trichoderma reesei as a model protein, we performed a mutagenesis study of the residues in proximity of the experimentally determined cleavage site which are possibly involved in the proteolytic process. However, the mutant forms of tyrosinase 2 were not secreted in a full-length form retaining the C-terminal domain, but they were processed to give a ~ 45 kDa active form. Aiming at explaining this phenomenon, we analysed in silico the properties of the C-terminal domain of tyrosinase 2, of 23 previously retrieved homologous tyrosinase sequences from fungi (C. Gasparetti, G. Faccio, M. Arvas, J. Buchert, M. Saloheimo, K. Kruus, Appl. Microbiol. Biotechnol. 86 (2010) 213–226) and of nine well-characterised polyphenol oxidases. Based on the results of our study, we exclude the key role of specific amino acids at the cleavage site in the proteolytic process and report an overall higher sensitivity to proteolysis of the linker region and of the whole C-terminal domain of fungal tyrosinases.

KW - c-terminal domain

KW - fungal tyrosinase

KW - protein processing

KW - proteolytic activation

KW - sensitivity to proteolysis

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DO - 10.1016/j.jinorgbio.2012.12.006

M3 - Article

VL - 121

SP - 37

EP - 45

JO - Journal of Inorganic Biochemistry

JF - Journal of Inorganic Biochemistry

SN - 0162-0134

ER -