A novel glycoprotein PLG-3 was obtained from the cultured mycelia of the edible mushroom Phellinus linteus AML1101 by isolating with cold water for 24 h and fractioned with DEAE-Sephrose Fast Flow column. It exhibited potent antiproliferative activity on A549 cells and HepG2 cells in vitro. After 24, 48, 72- h treatment with PLG-3, IC50 values of A549 cells and HepG2 cells were 45.77, 24.09, 16.54 μg/L and 47.78, 26.73, 18. μg/L, respectively. PLG-3 had a molecular weight of 2.5×104 Da, and consisted of approximately 85.5% of protein and 13.7% polysacchrides. It was composed of arabinose, xylose, mannose, glucose and galactose with a molar ratio of 1.46: 2.22: 7.74: 1.29:0.61.The obtained results proven that PLG-3 was a protein-dominant glycoprotein with singificant antiproliferative activity.