Optimization of different culture conditions for enhanced laccase production and its purification from Tricholoma giganteum AGHP

Abstract

Background

In current times, enzyme-catalyzed reactions have gained importance for the development of new chemical processes. These require the production of large quantity of enzyme at low cost. Solid-state fermentation (SSF) is an efficient process because this bioprocess has a potential to convert agro-industrial residues into valuable compounds. Hence, the current study focuses on the optimization of process parameters for the higher production of laccase using a novel basidiomycete fungi Tricholoma giganteum AGHP under solid-state fermentation (SSF). Further, the purification of laccase using column chromatographic technique was performed.

Results

Various physico-chemical parameters were evaluated and maximum production obtained was 2.69 × 10⁵ U/g using wheat straw as a dry substrate. Optimum pH was found to be 5.0 and the temperature of 30 °C with 0.3 mM copper as an inducer. The enzyme was purified from the initial protein preparation by two-step column chromatography. A yield of 10.49 % with 3.33-fold purification was obtained using Sephadex G-75 gel permeation chromatography. Further increase in purification (total) was found to be 10.80-fold with a yield of 8.50 % using DEAE Sephadex A-50 ion exchange column chromatography. The purified enzyme was identified as a monomeric protein with a molecular weight of 66 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE).

Conclusion

In view of the results obtained, we can conclude that the extracellular laccase production is governed by various cultural parameters such as pH, temperature, and the composition of culture medium. “One-factor-at-a-time” methodology was capable of establishing the optimum conditions that significantly increases the enzyme production several folds using lignocellulosic substrate. Therefore, laccase from T. giganteum AGHP has a potential in several industrial applications.

Keywords

Laccase / White rot fungi / Tricholoma giganteum / Solid-state fermentation / Inducers / Purification

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Funding

Department of Biotechnology, Ministry of Science and Technology(BT/PR5859/PID/6/696/2012)