Purification and characterization of two thermostable laccases from Pycnoporus sanguineus and potential role in degradation of endocrine disrupting chemicals uri icon


  • In this study, the purification and characterization of two thermostable laccases produced by Pycnoporus sanguineus CS43 (LacI and LacII) were performed. Also, their biotechnological potential was assessed through the degradation of endocrine disrupting chemicals (EDCs). Laccases were purified by ultrafiltration, ion exchange (IEX) and hydrophobic interaction chromatography achieving specific activities close to 285 U mg-1. The molecular weights of LacI and LacII, determined by SDS-electrophoresis, were 68 and 66 kDa, respectively. Both laccases showed high amino acid sequence similarity (91%) between them and high thermostability, at 50 and 60 °C (half-lives of 277.7 and 18 h for LacI, 35.8 and 2.25 h for LacII). The isoforms oxidized common laccase substrates such as 2,2¿-azino-bis (3-ethylbenzthiazoline-6- sulfonate (ABTS), 2,6-dimethoxyphenol (DMP) and guaiacol at acidic pH conditions. ABTS was the most efficient substrate, showing high specificity constants of 74,816 and 36,746 mM-1 s-1 for Lac I and LacII, and Michaelis constants (Km) of 6.9 and 12.2 ¿M respectively at pH 3. Both purified laccases remained active at high concentrations of organic solvents (acetonitrile, ethanol and acetone), with an IC50 (v/v) of >64%, 55% and 47% for LacI, and 33%, 52% and 31% for LacII, respectively. LacI and LacII were tested to degrade EDCs, nonylphenol and triclosan, with more than 95% removal after 8 h of treatment with 100 U/L at pH 5 as determined by means of HPLC. The high thermostability, unique Michaelis-Menten kinetic parameters, and organic solvent tolerance demonstrated for the isoforms produced by P. sanguineus CS43 render them promising candidates for industrial applications. LacI exerted a higher thermal and pH stability, tolerance against inhibitors and was a more efficient catalyst for ABTS and DMP than LacII. © 2014 The Authors.

Publication date

  • January 1, 2014