Phytocatalytic and cytotoxic activity of the purified laccase from bled resin of Pistacia atlantica Desf.

This study reports an efficient and fast procedure for the purification of laccase (PaL) obtained from the resin of Pistacia atlantica Desf. It was purified by one-step affinity chromatography and showed the specific activity of 393 U/mg with 81.9-fold purification. The molecular weight of PaL was estimated to be approximately 60 kDa using gel electrophoresis SDS-PAGE. Moreover, it depicted diphenolase activity and high affinity towards 2,6-dimethoxy phenol (K-m = 10.01 +/- 0.5 mM) and syringaldazine (K-m = 6.57 +/- 0.2 mM) comparing with plant-origin polyphenol oxidases reported in the literature. It should be noted that PaL possessed optimal activity at pH 7.5 and 45 degrees C. It also remained stable under different conditions of pH (6.5-8.0), temperature (25-45 degrees C), and when it was exposed to several metal ions. The MTT and flow cytometry assays demonstrated that the enzyme treatment significantly affected growth of HeLa, HepG2, and MDA-MB-231 cells with LC50 values of 4.83 +/- 0.02, 61 +/- 0.31, and 26.83 +/- 0.11 mu M after 72 h, respectively. Novelty statement: This is the first attempt to isolate and characterize a new oxidoreductase from the resin of Pistacia atlantica Desf., native species of Iran, to recruit it in cytotoxicity researches. In the purification process by an efficient affinity column (SBA-NH2-GA), the enzyme was eluted promptly with a satisfied yield. The purified laccase exerted higher affinity to diphenolic compounds and pH-thermal stability compared to other plant-derived polyphenol oxidases. The purified enzyme was found to show anti-oxidant capacity and significantly inhibited the growth of cancerous cells in vitro. PaL showed more cytotoxic activity towards HeLa and MDA-MB-231 cells by induction of apoptosis. The cytotoxic activity of the laccase was measured by flow cytometry. (c) 2021 Elsevier B.V. All rights reserved.

Automated summary

This study describes an efficient method for purifying laccase (PaL) from Pistacia atlantica Desf. resin, achieving a high specific activity and purification fold. PaL showed stability and high affinity towards specific substrates, as well as significant growth inhibition effects on cancer cells in vitro.