A ribonuclease with antimicrobial, antimitogenic and antiproliferative activities from the edible mushroom Pleurotus sajor-caju

Abstract

A 12 kDa ribonuclease preferential for poly U and with much lower activity toward poly A, poly G and poly C was isolated from fresh fruiting bodies of the mushroom Pleurotus sajor-caju. A purification procedure involving ion exchange chromatography on CM-cellulose, affinity chromatography on Red-Sepharose and Heparin-Sepharose, and fast protein liquid chromatography–gel filtration on Superdex 75 was used. The ribonuclease was adsorbed on all of the first three types of chromatographic media. It exhibited some activity toward herring sperm DNA and calf thymus DNA. The ribonuclease activity was unaffected in the presence of KCl (10 and 100 mM) and NaCl (100 mM and 1 M), but was strongly inhibited by CuSO 4 (0.01 and 0.1 mM) and less potently inhibited by other divalent salts including MgCl 2, CaCl 2, ZnCl 2, ZnSO 4 and FeSO 4. The optimal pH was 5.5 and the ribonuclease was stable up to 60 °C for 1 h. The ribonuclease inhibited mycelial growth in the fungi Fusarium oxysporum and Mycosphaerella arachidicola with an IC 50 value of 95 and 72 μM, respectively. Out of the 12 species of bacteria tested, only Pseudomonas aeruginosa and Staphylococcus aureus were inhibited in growth by the ribonuclease. Viability of the tumor cells HepG2 (hepatoma) and L1210 (leukemia) was reduced with an IC 50 of 0.22 and 0.1 μM, respectively in the presence of the ribonuclease. The ribonuclease inhibited translation in a cell-free rabbit reticulocyte lysate system with an IC 50 of 158 nM and 3 H -methyl-thymidine uptake by murine splenocytes with an IC 50 of 65 nM.