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Biochemical characterization of laccase from hairy root culture of Brassica juncea L. and role of redox mediators to enhance its potential for the decolorization of textile dyes

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Abstract

In vitro transgenic hairy root cultures provide a rapid system for physiological, biochemical studies and screening of plants for their phytoremediation potential. The hairy root cultures of Brassica juncea L. showed 92% decolorization of Methyl orange within 4 days. Out of the different redox mediators that were used to achieve enhanced decolorization, 2, 2′-Azinobis, 3-ethylbenzothiazoline-6-sulfonic acid (ABTS) was found to be the most efficient. Laccase activity of 4.5 U mg−1 of protein was observed in hairy root cultures of Brassica juncea L., after the decolorization of Methyl orange. Intracellular laccase produced by B. juncea root cultures grown in MS basal medium was purified up to 2.0 fold with 6.62 U mg−1 specific activity using anion-exchange chromatography. Molecular weight of the purified laccase was estimated to be 148 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The purified enzyme efficiently oxidized ABTS which was also required for oxidation of the other tested substrates. The pH and temperature optimum for laccase activity were 4.0 and 40°C, respectively. The purified enzyme was stable up to 50°C and was stable in the pH range of 4.0–6.0. Laccase activity was strongly inhibited by sodium azide, EDTA, dithiothreitol and l-cysteine. The purified enzyme decolorized various textile dyes in the presence of ABTS as an efficient redox mediator. These findings contribute to a better understanding of the enzymatic process involved in phytoremediation of textile dyes by using hairy roots.

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Abbreviations

SDS-PAGE:

Sodium dodecyl sulfate polyacrylamide gel electrophoresis

ABTS:

2,2′-Azinobis,3-ethylbenzothiazoline-6-sulfonic acid

MS:

Murashige and Skoog

ACS:

Acetosyringone

VAN:

Vanillin

SGZ:

Syringaldazine

HBT:

1-Hydroxybenzotriazole

SA:

Syringic acid

HQ:

Hydroquinone

DMP:

2,6-Dimethoxy phenol

PG:

Pyrogallol

DCIP:

2,6-Dichlorophenol indophenol

LMSs:

Laccase mediator systems

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Acknowledgments

VAB, AAT and UBJ wish to thank and express gratitude to Council of Scientific and Industrial Research (CSIR), New Delhi, India, for Emeritus Scientist, Senior Research Fellowship and Junior Research Fellowship, respectively. ANK wishes to thank the Department of Science and Technology (DST), New Delhi, India, for Junior Research Fellowship. SPG wishes to thank University Grants Commission, New Delhi for SAP-DRS-1 support.

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Authors and Affiliations

  1. Synthetic Biotechnology Laboratory, Division of Applied Life Sciences, Gyeongsang National University, Jinju, Korea Republic

    Amar A. Telke

  2. Department of Biochemistry, Shivaji University, Kolhapur, 416004, India

    Anuradha N. Kagalkar & Sanjay P. Govindwar

  3. Department of Biotechnology, Shivaji University, Kolhapur, 416004, India

    Umesh B. Jagtap & Vishwas A. Bapat

  4. Department of Biotechnology and Bioinformatics, Padmashree Dr. D.Y. Patil University, Sect-15/50 C. B. D. Belapur, Navi Mumbai, 400614, India

    Neetin S. Desai

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  1. Amar A. Telke
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  2. Anuradha N. Kagalkar
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  3. Umesh B. Jagtap
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  4. Neetin S. Desai
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  5. Vishwas A. Bapat
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  6. Sanjay P. Govindwar
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Correspondence to Sanjay P. Govindwar.

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Telke, A.A., Kagalkar, A.N., Jagtap, U.B. et al. Biochemical characterization of laccase from hairy root culture of Brassica juncea L. and role of redox mediators to enhance its potential for the decolorization of textile dyes. Planta 234, 1137–1149 (2011). https://doi.org/10.1007/s00425-011-1469-x

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