Lentinan is the main active substances in Lentinus edodes, but its structure is complex and the activity mechanism is not clear, so the application of lentinan is limited. Therefore, deepening the discussion of the structure-activity relationship of lentinan is helpful to fully explore the medicinal value of lentinan, and it is of great significance for the research and development of new products. Lentinus edodes crude polysaccharide was prepared by water extraction and alcohol precipitation method, and Lentinus edodes polysaccharide was deproteinized by Sevage method. EDTA-2Na was used to remove metal ions from lentina edodes. Congo red test, Fourier infrared spectroscopy, atomic force microscopy, DPPH, ABTS, hydroxyl radical scavenging experiments and antibacterial experiments were used to compare the changes of the structure, antioxidant and antibacterial activities of lentina edodes before and after metal ions removal. The results showed that after metal ions were removed, the rigidity of the triple helix structure of lentinus was weakened, and the chain thickness decreased from 1.2 mm to 0.8 mm. The scavenging ability of Lentinan on ABTS and hydroxyl free radicals decreased, while the scavenging ability of DPPH free radicals increased. When the concentration of polysaccharide was 1 mg/mL, the scavenging rates of DPPH, ABTS and hydroxylradical were 94.94%, 83.52% and 26.52%, respectively. The bacteriostatic test showed that the bacteriostatic effect of lentinan was decreased by removing metal ions. When the concentration of polysaccharide was 40 mg/mL, the range of inhibition zone against E. coli and Staphylococcus aureus decreased from 9.50 mm to 8.30 mm and 12.25 mm to 11.33 mm, respectively. Metal ions, as the main components of lentinan, participate in its structural composition and play an important role in its activity.