This work aimed to study and optimize the parameters affecting partition of a cellulase from the strain Bacillus velezensis A4 by aqueous two-phase system (ATPS). Four factors, molecular weight of polyethylene glycol (PEG), PEG concentration, phosphate salt concentration and pH, were investigated. A L-9(3(4)) orthogonal experiment was initially carried out to screen important factors affecting partitioning of the enzyme in ATPS. The detailed study of the parameters on enzyme partitioning in ATPS was carried out using Box-Behnken (Box-B) design of response surface methodology (RSM). The statistical analysis indicated the molecular weight of PEG, PEG concentration, and pH affect the recovery rate and purification factor of cellulase partitioning significantly. The model was built and analyzed through RSM analysis and validated by our experiment. The maximum recovery rate of enzyme (67.8%) and purification factor (1.14) could be achieved under the conditions of PEG 4000 (20.75%, w/w), K2HPO4 (8.5%, w/w), pH 8.5 in ATPS. The cellulase molecular weight was 35 kDa exhibited in zymogram and has an optimum temperature and pH at 60 degrees C and 5.0.