A novel hemagglutinin with antiproliferative activity against tumor cells from the hallucinogenic mushroom Boletus speciosus

Abstract

Little was known about bioactive compounds from the hallucinogenic mushroom Boletus speciosus. In the present study, a hemagglutinin (BSH, B. speciosus hemagglutinin) was isolated from its fruiting bodies and enzymatic properties were also tested. The chromatographic procedure utilized comprised anion exchange chromatography on Q-Sepharose, cation exchange chromatography on CM-Cellulose, cation exchange chromatography on SP-Sepharose, and gel filtration by FPLC on Superdex 75. The hemagglutinin was a homodimer which was estimated to be approximately 31 kDa in size. The activity of BSH was stable up to 60°C, while there was a precipitous drop in activity when the temperature was elevated to 70°C. BSH retained 25% hemagglutinating activity when exposed to 100 mM NaOH and 25 mM HCl. The activity was potently inhibited by 1.25 mM Hg(2+) and slightly inhibited by Fe(2+), Ca(2+), and Pb(2+). None of the sugars tested showed inhibition towards BSH. Its hemagglutinating activity towards human erythrocytes type A, type B, and type AB was higher than type O. The hemagglutinin showed antiproliferative activity towards hepatoma Hep G2 cells and mouse lymphocytic leukemia cells (L1210) in vitro, with IC50 of 4.7 μ M and 7.0 μ M, respectively. It also exhibited HIV-1 reverse transcriptase inhibitory activity with an IC50 of 7.1 μ M.

Figure 1

(a) Gel filtration of fraction on Superdex 75 HR 10/30 column by FPLC. Fraction SU1 represents purified hemagglutinin BSH. (b) SDS-PAGE of purified BSH. Right lane: molecular weight markers, from top downwards, phosphorylase b (94 kDa), bovine serum albumin (67 kDa), ovalbumin (43 kDa), carbonic anhydrase (30 kDa), soybean trypsin inhibitor (20 kDa), and lactalbumin (14.4 kDa).

Figure 2

Antiproliferative activities of BSH towards Hep G2 and L1210 cell lines in vitro. The IC₅₀ values towards Hep G2 and L1210 cells were 4.7 μM and 7.0 μM, respectively. Each value in both panels represents the means ± SD (n = 3). Different letters (a, b, c,…) next to the data points indicate statistically significant differences (P < 0.05) when the data were analyzed by analysis of variance followed by Duncan's multiple range tests.

Figure 3

Inhibitory activity of BSH towards HIV-1 reverse transcriptase. It exhibited HIV-1 RT inhibitory activity with an IC₅₀ of 7.1 μM. Each value in both panels represents the means ± SD (n = 3). Different letters (a, b, c,…) next to the data points indicate statistically significant differences (P < 0.05) when the data were analyzed by analysis of variance followed by Duncan's multiple range tests.