Characterization of a new tyrosinase from Pycnoporus species with high potential for food technological applications

S. Halaouli, Mi. Asther, Kristiina Kruus, L. Guo, M. Hamdi, J.-C. Sigoillot, M. Asther, A. Lomascolo (Corresponding Author)

Research output: Contribution to journalArticleScientificpeer-review

74 Citations (Scopus)

Abstract

Aims: Tyrosinase production by Pycnoporus cinnabarinus and Pycnoporus sanguineus was screened among 20 strains originating from various geographical areas, particularly from tropical environments. The tyrosinase from the most efficient strain was purified and characterized and tested for food additive applications.

Methods and Results: Monophenolase and diphenolase activities of tyrosinase were measured from cell lysate from the 20 Pycnoporus strains, for 8–10 days of cultivation. The strain P. sanguineus CBS 614.73 showed the highest productivity (45·4 and 163·6 U g−1 protein per day for monophenolase and diphenolase respectively). P. sanguineus CBS 614.73 tyrosinase was purified from concentrated cell lysate, anion‐exchange, size‐exclusion and hydroxyapatite chromatography, with a final yield of 2% and a purification factor of 35–38. The pure enzyme was a monomere with a molecular mass of 45 kDa and it showed four isoforms or isoenzymes with pI between 4·5–5. No N‐glycosylation was found. The N‐terminal amino acid sequence was IVTGPVGGQTEGAPAPNR. The enzyme was shown to be almost fully active in a pH range of 6–7, in a large temperature range (30–70°C), and was stable below 60°C. The main kinetic constants were determined. The tyrosinase was able to convert p‐tyrosol and p‐coumaric acid into hydroxytyrosol and caffeic acid, respectively, and it could also catalyse the cross‐linking formation of a model protein.

Conclusions: Among the genus Pycnoporus, known for the production of laccase, the strain P. sanguineus CBS 614.73 was shown to produce one other phenoloxidase, a new monomeric tyrosinase with a specific activity of 30 and 84 U mg−1 protein for monophenolase and diphenolase respectively.

Significance and Impact of the Study: This study identified P. sanguineus CBS 614.73 as a potential producer of a tyrosinase which demonstrated effectiveness in the synthesis of antioxidant molecules and in protein cross‐linking.

Original languageEnglish
Pages (from-to)332 - 343
Number of pages12
JournalJournal of Applied Microbiology
Volume98
Issue number2
DOIs
Publication statusPublished - 2005
MoE publication typeA1 Journal article-refereed

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Pycnoporus
Monophenol Monooxygenase
Food
Proteins
Laccase
Food Additives
Enzymes
Durapatite
Isoenzymes
Chromatography
Amino Acid Sequence
Protein Isoforms
Antioxidants

Keywords

  • antioxidants
  • cross-linking
  • polyphenol oxidase
  • Pycnoporus sanguineus
  • tyrosinase

Cite this

Halaouli, S. ; Asther, Mi. ; Kruus, Kristiina ; Guo, L. ; Hamdi, M. ; Sigoillot, J.-C. ; Asther, M. ; Lomascolo, A. / Characterization of a new tyrosinase from Pycnoporus species with high potential for food technological applications. In: Journal of Applied Microbiology. 2005 ; Vol. 98, No. 2. pp. 332 - 343.
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abstract = "Aims: Tyrosinase production by Pycnoporus cinnabarinus and Pycnoporus sanguineus was screened among 20 strains originating from various geographical areas, particularly from tropical environments. The tyrosinase from the most efficient strain was purified and characterized and tested for food additive applications. Methods and Results: Monophenolase and diphenolase activities of tyrosinase were measured from cell lysate from the 20 Pycnoporus strains, for 8–10 days of cultivation. The strain P. sanguineus CBS 614.73 showed the highest productivity (45·4 and 163·6 U g−1 protein per day for monophenolase and diphenolase respectively). P. sanguineus CBS 614.73 tyrosinase was purified from concentrated cell lysate, anion‐exchange, size‐exclusion and hydroxyapatite chromatography, with a final yield of 2{\%} and a purification factor of 35–38. The pure enzyme was a monomere with a molecular mass of 45 kDa and it showed four isoforms or isoenzymes with pI between 4·5–5. No N‐glycosylation was found. The N‐terminal amino acid sequence was IVTGPVGGQTEGAPAPNR. The enzyme was shown to be almost fully active in a pH range of 6–7, in a large temperature range (30–70°C), and was stable below 60°C. The main kinetic constants were determined. The tyrosinase was able to convert p‐tyrosol and p‐coumaric acid into hydroxytyrosol and caffeic acid, respectively, and it could also catalyse the cross‐linking formation of a model protein. Conclusions: Among the genus Pycnoporus, known for the production of laccase, the strain P. sanguineus CBS 614.73 was shown to produce one other phenoloxidase, a new monomeric tyrosinase with a specific activity of 30 and 84 U mg−1 protein for monophenolase and diphenolase respectively. Significance and Impact of the Study: This study identified P. sanguineus CBS 614.73 as a potential producer of a tyrosinase which demonstrated effectiveness in the synthesis of antioxidant molecules and in protein cross‐linking.",
keywords = "antioxidants, cross-linking, polyphenol oxidase, Pycnoporus sanguineus, tyrosinase",
author = "S. Halaouli and Mi. Asther and Kristiina Kruus and L. Guo and M. Hamdi and J.-C. Sigoillot and M. Asther and A. Lomascolo",
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Characterization of a new tyrosinase from Pycnoporus species with high potential for food technological applications. / Halaouli, S.; Asther, Mi.; Kruus, Kristiina; Guo, L.; Hamdi, M.; Sigoillot, J.-C.; Asther, M.; Lomascolo, A. (Corresponding Author).

In: Journal of Applied Microbiology, Vol. 98, No. 2, 2005, p. 332 - 343.

Research output: Contribution to journalArticleScientificpeer-review

TY - JOUR

T1 - Characterization of a new tyrosinase from Pycnoporus species with high potential for food technological applications

AU - Halaouli, S.

AU - Asther, Mi.

AU - Kruus, Kristiina

AU - Guo, L.

AU - Hamdi, M.

AU - Sigoillot, J.-C.

AU - Asther, M.

AU - Lomascolo, A.

PY - 2005

Y1 - 2005

N2 - Aims: Tyrosinase production by Pycnoporus cinnabarinus and Pycnoporus sanguineus was screened among 20 strains originating from various geographical areas, particularly from tropical environments. The tyrosinase from the most efficient strain was purified and characterized and tested for food additive applications. Methods and Results: Monophenolase and diphenolase activities of tyrosinase were measured from cell lysate from the 20 Pycnoporus strains, for 8–10 days of cultivation. The strain P. sanguineus CBS 614.73 showed the highest productivity (45·4 and 163·6 U g−1 protein per day for monophenolase and diphenolase respectively). P. sanguineus CBS 614.73 tyrosinase was purified from concentrated cell lysate, anion‐exchange, size‐exclusion and hydroxyapatite chromatography, with a final yield of 2% and a purification factor of 35–38. The pure enzyme was a monomere with a molecular mass of 45 kDa and it showed four isoforms or isoenzymes with pI between 4·5–5. No N‐glycosylation was found. The N‐terminal amino acid sequence was IVTGPVGGQTEGAPAPNR. The enzyme was shown to be almost fully active in a pH range of 6–7, in a large temperature range (30–70°C), and was stable below 60°C. The main kinetic constants were determined. The tyrosinase was able to convert p‐tyrosol and p‐coumaric acid into hydroxytyrosol and caffeic acid, respectively, and it could also catalyse the cross‐linking formation of a model protein. Conclusions: Among the genus Pycnoporus, known for the production of laccase, the strain P. sanguineus CBS 614.73 was shown to produce one other phenoloxidase, a new monomeric tyrosinase with a specific activity of 30 and 84 U mg−1 protein for monophenolase and diphenolase respectively. Significance and Impact of the Study: This study identified P. sanguineus CBS 614.73 as a potential producer of a tyrosinase which demonstrated effectiveness in the synthesis of antioxidant molecules and in protein cross‐linking.

AB - Aims: Tyrosinase production by Pycnoporus cinnabarinus and Pycnoporus sanguineus was screened among 20 strains originating from various geographical areas, particularly from tropical environments. The tyrosinase from the most efficient strain was purified and characterized and tested for food additive applications. Methods and Results: Monophenolase and diphenolase activities of tyrosinase were measured from cell lysate from the 20 Pycnoporus strains, for 8–10 days of cultivation. The strain P. sanguineus CBS 614.73 showed the highest productivity (45·4 and 163·6 U g−1 protein per day for monophenolase and diphenolase respectively). P. sanguineus CBS 614.73 tyrosinase was purified from concentrated cell lysate, anion‐exchange, size‐exclusion and hydroxyapatite chromatography, with a final yield of 2% and a purification factor of 35–38. The pure enzyme was a monomere with a molecular mass of 45 kDa and it showed four isoforms or isoenzymes with pI between 4·5–5. No N‐glycosylation was found. The N‐terminal amino acid sequence was IVTGPVGGQTEGAPAPNR. The enzyme was shown to be almost fully active in a pH range of 6–7, in a large temperature range (30–70°C), and was stable below 60°C. The main kinetic constants were determined. The tyrosinase was able to convert p‐tyrosol and p‐coumaric acid into hydroxytyrosol and caffeic acid, respectively, and it could also catalyse the cross‐linking formation of a model protein. Conclusions: Among the genus Pycnoporus, known for the production of laccase, the strain P. sanguineus CBS 614.73 was shown to produce one other phenoloxidase, a new monomeric tyrosinase with a specific activity of 30 and 84 U mg−1 protein for monophenolase and diphenolase respectively. Significance and Impact of the Study: This study identified P. sanguineus CBS 614.73 as a potential producer of a tyrosinase which demonstrated effectiveness in the synthesis of antioxidant molecules and in protein cross‐linking.

KW - antioxidants

KW - cross-linking

KW - polyphenol oxidase

KW - Pycnoporus sanguineus

KW - tyrosinase

U2 - 10.1111/j.1365-2672.2004.02481.x

DO - 10.1111/j.1365-2672.2004.02481.x

M3 - Article

VL - 98

SP - 332

EP - 343

JO - Journal of Applied Microbiology

JF - Journal of Applied Microbiology

SN - 1364-5072

IS - 2

ER -