Using the high efficient copper-adsorbing yeast strain Y17 as absorbing material, the major affect factors including pH, original concentration of Cu2+, cell biomass, adsorption time and temperature were examined, and then the absorbing sites of the Y17 was determined. The results showed that the solution pH was the most dominate factor which affected the biosorption of Cu2+, the other affecting factors were the initial concentration of Cu2+, the cell biomass added, and adsorption time, respectively; the temperature had little effect on the rate of biosorption. The orthogonal experiment showed that the optimal absorption condition was as follow: the solution pH was 5.0, the absorption time was 40 min, the cell biomass of Y17 added was 5.0 g/L, and the concentration of Cu2+ was 8 mmol/L; the highest adsorbing rate was up to 82.7% at this condition. Based on the results of different pretreatments and the desorption of Cu2+, the cell wall of Y17 was identified as the main place occurring boisorption process, and the -NH2 group, -COOH group on the surface of the yeast cells played an important role on the boisorption process.