Abstract
Laccases are diphenol oxidases that have numerous applications to biotechnological processes. In this study, the laccase was produced from the thermophilic actinomycetes, Thermobifida fusca BCRC 19214. After 36 h of fermentation in a 5-liter fermentor, the culture broth accumulated 4.96 U/ml laccase activity. The laccase was purified 4.64-fold as measured by specific activity from crude culture filtrate by ultrafiltration concentration, Q-Sepharose FF and Sephacryl™ S-200 column chromatography. The overall yield of the purified enzyme was 7.49%. The molecular mass of purified enzyme as estimated by SDS-PAGE and by gel filtration on Sephacryl™ S-200 was found to be 73.3 kDa and 24.7 kDa, respectively, indicating that the laccase from T. fusca BCRC 19214 is a trimer. The internal amino acid sequences of the purified laccase, as determined by LC-MS/MS, had high homology with a superoxide dismutase from T. fusca YX. Approximately 95% of the original activity remained after treatment at 50°C for 3 h. and approximately 75% of the original activity remained after treatment at pH 10.0 for 24 h. This laccase could oxidize dye intermediates, especially 2,6-dimethylphenylalanine and p-aminophenol, to produce coloring. This is the first report on laccase properties from thermophilic actinomycetes. These properties suggest that this newly isolated laccase has potential for specific industrial applications.
Highlights
Laccases (E.C. 1.10.3.2) are well-known enzymes that were first isolated from the lacquer tree, Rhus vernicifera
Production of the laccase in fermentor Bagasse, corncob and pine sawdust were used as carbon sources to produce the laccase from T. fusca BCRC 19214
The highest extracellular laccase activity was observed when T. fusca BCRC 19214 was grown in a mineral medium containing bagasse as the carbon source
Summary
Laccases (E.C. 1.10.3.2) are well-known enzymes that were first isolated from the lacquer tree, Rhus vernicifera. They have received increasing attention in recent decades due to their ability to oxidize both phenolic and nonphenolic lignin-related compounds and highly recalcitrant environmental pollutants, which makes them very useful for applications related to biotechnological processes (Mukhopadhyay et al, 2013; Couto and Herrera, 2006; Albino et al, 2004). In a typical hair color product, the dye intermediates are p-diamines and p-aminophenols, and hydrogen peroxide is used as the oxidant in the dyeing process. After mixing, they form chromatic indo dyes at the time of use. Laccase-based hair dyes are less irritating and easier to handle than current hair dyes, as laccases replaced hydrogen peroxide as the oxidizing agent in the dyeing formula (Couto and Herrera, 2006)
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