Phytocatalytic and Cytotoxic Activity of the Purified Laccase From Bled Resin of Pistacia Atlantica Desf. Publisher Pubmed



Elyasighahfarokhi A^{1, 2} ; Hashemi S² ; Saeedi M^{3, 4} ; Khanavi M^{1, 5} ; Faramarzi MA² Show Affiliations
Source: International Journal of Biological Macromolecules Published:2021

Abstract

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 (Km = 10.01 ± 0.5 mM) and syringaldazine (Km = 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 °C. It also remained stable under different conditions of pH (6.5–8.0), temperature (25–45 °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 μM 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. © 2021 Elsevier B.V.